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)

The effects of erythromycin on the NADPH oxidase activity in the neutrophils of normal subjects and patients with bronchiolitis were investigated. In the patients receiving erythromycin, NADPH oxidase activity was significantly lower in a whole-cell system than that before therapy. Erythromycin was also found to inhibit the superoxide generation of neutrophils exposed to phorbol myristate acetate in a whole-cell system and the activation of superoxide-generating NADPH oxidase by sodium lauryl sulfate (sodium dodecyl sulfate) in a cell-free system. The concentration of the drug required for 50 percent inhibition of the oxidase was 0.7 mM in the whole-cell system and 0.2 mM in the cell-free system. These results suggest that erythromycin, an antibiotic which penetrates well into human neutrophils, may exhibit an anti-inflammatory action due to inhibiting of neutrophil NADPH oxidase activation.
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PMID:Anti-inflammatory action of erythromycin. Its inhibitory effect on neutrophil NADPH oxidase activity. 840 90

To combat bacterial infection, phagocytes generate superoxide (O2-) and other microbicidal oxygen radicals. NADPH oxidase, the enzyme responsible for O2- synthesis, is deficient in chronic granulomatous disease (CGD) patients. Although O2- generation is accompanied by a large burst of metabolic acid production, intracellular pH (pHi) remains near neutrality due to the concomitant stimulation of H+ extrusion. Three major pathways contribute to pHi regulation in activated phagocytes: Na+/H+ exchange, vacuolar-type H+ pumps, and a H+ conductance. The present study analyzed the relationship between activation of the NADPH oxidase and stimulation of the H+ extrusion mechanisms in human blood neutrophils. Phorbol ester-induced activation of Na+/H+ exchange and H+ pumping occurred normally in cells from CGD patients. Unlike normal individuals, however, CGD patients were unable to activate the H+ conductive pathway. Thus, activation of the H+ conductance appears to be contingent on the assembly of a functional NADPH oxidase. These findings imply a dual role of the NADPH oxidase in O2- synthesis and in the regulation of pHi. The oxidase (or some components thereof) may itself undertake H+ translocation or, alternatively, may signal the activation of a separate H+ conducting entity.
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PMID:Abnormal activation of H+ conductance in NADPH oxidase-defective neutrophils. 842 13

A flavoprotein dehydrogenase assayed for the activity of electron transfer from NADPH to cytochrome c was highly purified from the cytosolic fraction of differentiated human promyelocytic leukemia HL-60 cells. The purified enzyme had an apparent molecular mass of 68 kDa by sodium dodecyl sulfate gel electrophoresis and an equimolar amounts of flavin mononucleotide and flavin-adenine dinucleotide. The purification factor of the enzyme with respect to the cytosolic fraction was close to 1100 and the recovery of activity was approximately 18%. Reduction of cytochrome c by NADPH indicated Michaelis-Menten kinetics with a Km value of 1.50 microM for NADPH. When cytochrome c was the varied substrate, a Km value of 4.10 microM was obtained. NADH was not an effective electron donor for cytochrome c reduction and NADPH-dependent reduction of nitroblue tetrazolium was negligibly small. The purified enzyme alone did not exhibit superoxide production, and NADPH oxidase activity was not markedly stimulated upon incubation of the reductase with cytochrome b558 purified from porcine neutrophils. The purified flavoprotein gave a positive cross-reactivity to polyclonal antibodies raised to microsomal NADPH-cytochrome P450 reductase, indicating structural homology between these enzymes. The catalytic properties of the purified NADPH-cytochrome c reductase have similarities to those of liver NADPH-cytochrome P450 reductase.
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PMID:Characterization of superoxide dismutase-insensitive cytochrome c reductase activity in HL-60 cytosol as NADPH-cytochrome P450 reductase. 848 36

We studied the effects of antiallergic drugs, epinastine, ketotifen, oxatomide, mequitazine and cromolyn sodium on superoxide anion (O2-) generation from rat neutrophils. Epinastine, ketotifen, oxatomide and mequitazine dose-dependently prevented the N-formyl-Met-Leu-Phe- and phorbol 12-myristate 13-acetate-induced O2- generation, but cromolyn sodium did not prevent it. When membrane and cytosol fractions were incubated with each drug, epinastine, ketotifen and mequitazine prevented O2- generation. On the other hand, when only the membrane fraction was incubated with each drug, ketotifen and mequitazine prevented O2- generation, but epinastine did not. Epinastine may inhibit the NADPH oxidase system through the obstruction of NADPH oxidase-associated cytosol components.
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PMID:Inhibitory effect of epinastine on superoxide generation by rat neutrophils. 853 20

The effect of the S-nitrosothiol (RSNO) on the activation of NADPH oxidase in human neutrophils was studied using an in vitro translocation system in which an anionic amphiphil, such as sodium dodecyl sulfate or arachidonate, plays a role as an activator. When membranes pretreated with RSNO and a cytosol fraction from resting neutrophils were combined to reconstitute the NADPH oxidase, both translocation of the cytosolic NADPH oxidase components such as p47phox and p67phox to the plasma membrane fraction and subsequent superoxide generation was inhibited. However, RSNO had no effect on O2- production when added after enzyme activation. A similar inhibition of translocation of recombinant p47phox was observed with RSNO-treated membrane. When the RSNO-treated membrane fraction was exposed to 2-mercaptoethanol the inhibition was reversed. The data suggest that RSNO inhibits translocation of p47phox or p47phox containing cytosolic complex via a direct effect on the membrane component of the NADPH oxidase.
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PMID:Attenuation of p47phox and p67phox membrane translocation as the inhibitory mechanism of S-nitrosothiol on the respiratory burst oxidase in human neutrophils. 860 52

The regulation of the intracelluar pH (pHi) during spreading of human neutrophils was studied by a combination of fluorescence imaging and video microscopy. Spreading on adhesive substrates caused a rapid and sustained cytosolic alkalinization. This pHi increase was prevented by the omission of external Na+, suggesting that it results from the activation of Na+/H+ exchange. Spreading-induced alkalinization was also precluded by the compound HOE 694 at concentrations that selectively block the NHE-1 isoform of the Na+H+ antiporter. Inhibition of Na+/H+ exchange by either procedure unmasked a sizable cytosolic acidification upon spreading, indicative of intracellular acid production. The excess acid generation was caused, at least in part, by the activation of the respiratory burst, since the acidification closely correlated with superoxide production, measured in single spreading neutrophils with dihydrorhodamine-123, and little acid production was observed in the presence of diphenylene iodonium, a blocker of the NADPH oxidase. Moreover, neutrophils from chronic granulomatous disease patients, which do not produce superoxide, failed to acidify. Comparable pHi changes were observed when beta 2 integrins were selectively activated during spreading on surfaces coated with anti-CD18 antibodies. When integrin engagement was precluded by pretreatment with soluble anti-CD18 antibody, the pHi changes associated with spreading on fibrinogen were markedly reduced. Inhibition of microfilament assembly with cytochalasin D precluded spreading and concomitantly abolished superoxide production and the associated pHi changes, indicating that cytoskeletal reorganization and/or an increase in the number of adherence receptors engaged are required for the responses. Neutrophils spread normally when the oxidase was blocked or when pHi was clamped near physiological values with nigericin. Spreading, however, was strongly inhibited when pHi was clamped at acidic values. Our results indicate that neutrophils release superoxide upon spreading, generating a burst of intracellular acid production. The concomitant activation of the Na+/H+ antiport not only prevents the deleterious effects of the acid released by the NADPH oxidase, but induces a net cytosolic alkalinization. Since several functions of neutrophils are inhibited at an acidic pHi, the coordinated activation of pHi regulatory mechanisms along with the oxidase is essential for sustained microbicidal activity.
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PMID:Intracellular pH regulation during spreading of human neutrophils. 868 73

Upon stimulation, inactive subunits of monocyte NADPH oxidase (NOX) are assembled in the membrane to generate the active enzyme responsible for oxidative burst. Phosphorylation of the 47 kDa NOX cytoplasmic subunit (47 kDa band) by protein kinase C (PKC) is important for NOX assembly and activation. Alternatively, NOX is activated in vitro by sodium dodecyl sulfate (SDS) or amphiphiles via a phosphorylation-independent mechanism. Previous data indicate that phagocytosis of malarial pigment hemozoin inhibits oxidative burst and PKC activity (Schwarzer, E., Turrini, F., Giribaldi, G., Cappadoro, M. and Arese, P. (1993) Biochim. Biophys. Acta, 1181, 51-54). We show here that SDS-stimulated NOX activity and phorbol 12-myristate 13-acetate (PMA)-induced oxidative burst dropped by 54% and 46% of control values 2 h after hemozoin phagocytosis, respectively. SDS-stimulated NOX activity remained roughly constant until 12 h, whereas oxidative burst dropped further by approx. 60% and 75% of control values 6 h and 12 h after hemozoin phagocytosis. Reconstitution experiments indicate that damage was localized to cytosolic NOX subunit(s). Membrane assembly of active NOX was defective in PMA-(PKC-dependent stimulation) and FMLP-(PKC-dependent and independent stimulation) stimulated hemozoin-fed monocytes. Labeling experiments with [32P]orthophosphate or [gamma-32P]ATP showed that endogenous PKC-dependent phosphorylation of the 47 kDa band was unaffected 12 h and impaired only 24 h after hemozoin phagocytosis. Thus, only long-term inhibition of NOX may additionally depend on superimposed PKC inhibition.
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PMID:Phagocytosis of malarial pigment hemozoin inhibits NADPH-oxidase activity in human monocyte-derived macrophages. 878 35

Hydrogen peroxide injected into the inflow cannula of isolated ventilated rat lungs produced a dose-dependent vasoconstriction in the range 0.25-10 mM, with maximum response between 2-5 mM. The effects of H2O2 can be influenced by ionophores or specific inhibitors of ionic channels or pumps. A key role is played by sodium ions which govern the subsequent inflow or outflow of calcium, an ion mediating the vasoconstriction. A physiological role for H2O2 generated by NADPH oxidase is postulated.
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PMID:Role of ion fluxes in hydrogen peroxide pulmonary vasoconstriction. 878 97

The NADPH oxidase complex of activated neutrophils consists of a membrane-bound flavocytochrome b and cytosolic activation factors. Despite its ability to react with O2, the heme b component of the flavocytochrome is insensitive to cyanide and CO2, and slowly reactive to butyl isocyanide. We report here that arachidonic acid, an anionic amphophil which elicits oxidase activation in a cell-free system induces a transition of the heme iron of the neutrophil flavocytochrome b from a low-spin hexacoordinated state to a high-spin pentacoordinated state and promotes the binding of butyl isocyanide to the heme b. Low-temperature EPR spectra of air-oxidized flavocytochrome b either purified or in its membrane-bound form showed a low-spin signal at g = 3.26 and a high-spin signal at g = 6.0. Upon addition of arachidonic acid, the g = 3.26 signal vanished; a low-spin signal at g = 2.23 appeared, and the signal at g = 6.0 progressively increased. The subsequent addition of butyl isocyanide resulted in the decrease of the g = 6.0 and g = 2.23 signals and in the appearance of a new low-spin signal at g = 2.33. Consistent with the EPR results, upon addition of arachidonic acid to oxidized flavocytochrome b, a 2.5 nm blue shift of the Soret peak was detected in low-temperature optical spectra. The subsequent addition of butyl isocyanide resulted in the emergence of a peak at 432 nm reflecting the formation of a butyl isocyanide-oxidized heme b complex. In the case of sodium dithionite-reduced flavocytochrome b, arachidonic acid promoted the binding of butyl isocyanide to the reduced heme b, as shown by the emergence of a peak at 434 nm and the decrease of the alpha band at 558 nm. The same promoting effect was encountered with sodium dodecyl sulfate, an anionic amphophil capable of eliciting oxidase activation like arachidonic acid. In contrast to arachidonic acid, arachidonic acid methyl ester was ineffective and counteracted the effect of arachidonic acid. Butyl isocyanide added to intact neutrophils was found to bind to heme b, only after the cells have been activated. These data demonstrate the transient accumulation of a pentacoordinated form of the heme iron of flavocytochrome b under in vitro and in vivo conditions; the pentacoordinated form of the reduced heme b is postulated to react with O2 to generate the superoxide anion.
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PMID:Electron transfer across the O2- generating flavocytochrome b of neutrophils. Evidence for a transition from a low-spin state to a high-spin state of the heme iron component. 887 8

Mechanisms for the cell-free activation of NADPH oxidase by sodium dodecyl sulfate (SDS) and arachidonate were compared in relation to their responsiveness to short chain diacylglycerols. The plasma membrane and cytosol prepared from guinea pig neutrophils were used for the cell-free system. The activation of NADPH oxidase by SDS was enhanced about 5- to 10-fold by 1,2-dioctanoylglycerol (diC8), but not by either 1,2-dihexanoylglycerol (diC6) or 1,2-didecanoylglycerol (diC10). However, none of these diacylglycerols potentiated the NADPH oxidase activation by arachidonate. The maximal extent of activation by the combination of SDS and diC8 was similar to that by arachidonate alone. In the presence of sufficient amounts of diC8 and SDS, GTP gamma S potentiated the activation of NADPH oxidase. The potentiating activity of diC8 was preserved in the membrane fraction, not in the cytosol fraction. These results suggest that arachidonate may possess the functions of both SDS and diC8 in the activation. In addition, diC8 and GTP gamma S seem to independently enhance the NADPH oxidase activation.
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PMID:Differences in mechanisms for cell-free NADPH oxidase activation between arachidonate and sodium dodecyl sulfate. 888 29


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