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:1.6.99.6 (NADPH oxidase)
10,295 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The effects of anti-inflammatory drugs (acetylsalicylic acid, ASA; salicylic acid, SA; indomethacin, IM; hydrocortisone, HC) on the respiratory burst oxidase (NADPH oxidase) from human neutrophils in both whole cell and fully soluble (cell-free) systems were investigated. These drugs were found to inhibit the superoxide generation by human neutrophils exposed to phorbol myristate acetate in a whole cell system and the activation of superoxide-generating NADPH oxidase by sodium dodecyl sulfate in cell-free systems. Concentrations of these drugs required for 50% inhibition of the oxidase (ID50) were; ASA (more than 3.0 mM in the whole cell system and 1.35 mM in the cell-free system), SA (more than 3.0 mM in the whole cell system and 1.30 mM in the cell-free system), IM (180 microM in both systems) and HC (50 microM in the whole cell system and 40 microM in the cell-free system). In addition, these drugs time-dependently inhibited the activation of NADPH oxidase in cell-free systems. In the cell-free system, all of the drugs did not change the Km values for NADPH of the oxidase. These results suggest that these anti-inflammatory drugs, especially HC and IM, inhibit the reconstitution (activation) of neutrophil NADPH oxidase enzyme in the cell-free (whole cell) system.
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PMID:[Studies on relationships between the inhibition of human neutrophil NADPH oxidase by anti-inflammatory drugs and development of bacterial infections]. 224 89

Rifapentine (R773, DL473) is a long-acting antituberculous drug used in China. In our experiments we have found some manifestations of induction of hepatic mixed function oxidase system in mice following pretreatment with rifapentine or phenobarbital. Both rifapentine and phenobarbital significantly increased the rate of antipyrine and pentobarbital metabolism in vivo. They also increased liver weight, the content of liver microsomal protein and cytochrome P-450, the activity of NADPH-cytochrome C reductase and NADPH oxidase. SDS-polyacylamide gel electrophoresis showed that the relative proportions of some polypeptide bands in mice microsomal fraction were significantly changed following rifapentine or phenobarbital pretreatment. The results indicate that rifapentine, like phenobarbital, is a potent inducer of hepatic mixed function oxidase system in mice and that it should be used carefully in clinical therapy, when combined with other drugs.
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PMID:Inductive effects of rifapentine on mice hepatic mixed function oxidase system. 231 33

Kinetics of activation of the NADPH oxidase in a fully soluble cell-free system from phorbol myristate acetate (PMA)-stimulated human neutrophils were investigated. In a cell-free system in which Mg2+ and sodium dodecyl sulfate, an anionic detergent required for the activation of NADPH oxidase are contained, cytosol prepared from PMA-stimulated neutrophils failed to activate PMA-stimulated neutrophil oxidase. However, cytosol prepared from resting (control) neutrophils was capable of activating PMA-stimulated neutrophil oxidase in a cell-free system in which its Km for NADPH was almost similar to that of control neutrophil oxidase. Cytosol from PMA-stimulated neutrophils could not activate control neutrophil oxidase, although it did not contain any inhibitors of NADPH oxidase activation. These results suggest that, in PMA-stimulated neutrophils, cytosolic activation factors may be consumed or exhausted, and that the affinity for NADPH of PMA-stimulated neutrophil oxidase may be the same as that of control neutrophil oxidase.
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PMID:Activation of the NADPH oxidase in a cell-free system from human neutrophils stimulated by phorbol myristate acetate. 234

1. A microtechnique for quantitating human neutrophil NADPH oxidase in a cell-free system is described. 2. This spectrophotometric discontinuous (fixed time) method is less material-consuming than existing methods and is more useful for experiments in which superoxide production by neutrophils must be measured in a large number of samples. 3. Measurement of NADPH oxidase using the new method can be accomplished in a final vol of 0.15 ml. 4. In the assay, neutrophil membranes solubilized with deoxycholate were incubated for 3 min with cytosolic fractions, magnesium, sodium dodecyl sulfate, and cytochrome c in the absence of NADPH to preincubate the oxidase before the addition of the reducing agent. 5. The reaction was started by adding NADPH and 2 min later terminated by adding superoxide dismutase. 6. The apparent Km for NADPH obtained by the new method was almost the same as that by the authorized method (39.2 +/- 3.1 SD vs 36.8 +/- 1.6). Activation of neutrophil NADPH oxidase was characterized using the new assay method.
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PMID:A microtechnique for neutrophil respiratory burst oxidase in a cell-free system--characterization of oxidase activation system. 239 Aug 59

A histochemical analysis of reaction rates of a series of enzymes was performed in electromotor neurons of the weakly electric fish Apteronotus leptorhynchus. These neurons were selected because of their functional homogeneity. The high metabolic activity of these cells as well as their large size facilitate cytophotometric analysis in cryostat sections. Sections were incubated for the activity of hexokinase, glucose-6-phosphate dehydrogenase, succinate dehydrogenase, NADPH dehydrogenase, NADPH ferrihaemoprotein reductase and beta-hydroxybutyrate dehydrogenase. All media contained polyvinyl alcohol as tissue stabilizer and Nitro BT as final electron acceptor. Measurements were performed with a Vickers M85a cytophotometer. Linear relationships between the specific formation of formazan (test minus control reaction) and incubation time were obtained for all enzymes although some reactions showed an initial lag phase or an intercept with the ordinate. The relatively high activities of hexokinase, succinate dehydrogenase and the extremely low activity of hydroxybutyrate dehydrogenase indicate that energy is mainly supplied by glycolysis. Glucose-6-phosphate dehydrogenase showed a high activity whereas NADPH reductase and dehydrogenase activity were low in electromotor neurons, indicating that the NADPH generated is largely used for biosynthesis. Despite their synchronous firing pattern activity, electromotor neurons showed a considerable heterogeneity with respect to their metabolic activity.
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PMID:Enzyme reaction rate studies in electromotor neurons of the weakly electric fish Apteronotus leptorhynchus. 251 71

Lipopolysaccharide (LPS) pretreatment "primes" neutrophils to release increased amounts of superoxide anion (O2-) when stimulated. We investigated the molecular basis of this enhanced activity. Comparison of kinetic parameters of the respiratory burst NADPH oxidase in unstimulated LPS-primed and control neutrophils disclosed a similar Km for NADPH and no difference was seen in the content of cytochrome b. Pertussis toxin, which inhibits some G proteins, did not prevent priming. Change in membrane potential (delta psi) was five-fold greater in LPS-primed cells and paralleled the increased O2- release. Cytofluorographic analysis indicated that the increased change in delta psi was due to the creation of a new population of active cells. Changes in the concentration of intracellular free Ca2+ ([Ca2+]i) are believed to antecede changes in delta psi. There was a consistent increment (67 +/- 8%, n = 12) in resting [Ca2+]i in cells preincubated with LPS compared with control. When stimulated, the peak [Ca2+]i was significantly higher in LPS-primed cells. Ca2+-dependent protein kinase C activity was unaltered in resting and FMLP-stimulated neutrophils preexposed to LPS. Addition to cells of the intracellular Ca2+ chelator MAPTAM before preincubation with LPS blocked the changes in [Ca2+]i and the enhanced respiratory burst that characterize LPS priming. The increased resting [Ca2+]i in LPS-primed cells may enhance stimulus-induced cellular activity by modifying a Ca2+-dependent step in signal transduction.
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PMID:Lipopolysaccharide priming of human neutrophils for an enhanced respiratory burst. Role of intracellular free calcium. 253 46

Ebselen (PZ51, 2-phenyl-1,2-benzoisoselenazol-3-(2H)-one) was shown to be an inhibitor of human granulocyte oxidative burst stimulated by phorbol myristate acetate (IC50 25 microM). Estimation of the primary oxygen metabolites of the burst was complicated by the redox chemistry of Ebselen. Ebselen inhibited NADPH-stimulated superoxide generation by a partially purified NADPH oxidase preparation with an IC50 of 0.5-1.0 microM. Ebselen was also shown to inhibit the activity of partially purified Ca2+- and phospholipid-dependent protein kinase C (IC50 ca. 0.5 microM). Phorbol ester-stimulated phosphorylation of protein in intact cells was inhibited by Ebselen (IC50 ca. 50 microM). These pharmacodynamic properties of Ebselen are discussed in terms of its anti-inflammatory activity in vivo. The findings are also discussed in terms of Ebselen's known ability to interact with sulfhydryl components of cells, particularly critical thiol components of the enzymes studied.
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PMID:Studies on the anti-inflammatory activity of ebselen. Ebselen interferes with granulocyte oxidative burst by dual inhibition of NADPH oxidase and protein kinase C? 253 84

A kinetic model was developed to describe the production of superoxide (O2-) by single pulmonary alveolar macrophages (PAM). Model predictions were compared with experimental results obtained from single rat PAM. The O2- was quantified by measuring the reduction of nitro blue tetrazolium (NBT) to a diformazan precipitate (NBTH2) from video-recorded images of individual cells. The kinetic model considered three reactions: 1) the production of extracellular O2- from the reduction of oxygen by NADPH oxidase using intracellular NADPH as the substrate, 2) the subsequent dismutation of O2- to form H2O2, and 3) the reaction of O2- and NBT to form diformazan. NBT specificity for O2- was analyzed by comparing results in the presence and absence of superoxide dismutase (SOD) that catalyzes the dismutation of O2- to H2O2. Measured PAM heterogeneity was accounted for by varying the concentration of intracellular NADPH, its rate of depletion, and the concentration of intracellular NADPH oxidase in the kinetic model. Model predictions compared favorably with experimental results except when SOD was present. This discrepancy may be due to diffusional limitations because NBT is a relatively small molecule (818 mol wt) compared with SOD (34,000 mol wt). In addition, the cell surface is both ruffled and negatively charged, which may introduce steric hindrances and/or electrostatic effects, since SOD is also negatively charged, whereas NBT is positively charged.
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PMID:A kinetic model of superoxide production from single pulmonary alveolar macrophages. 253 74

In neutrophils of patients with essential hypertension the NADPH-dependent O2- production elicited by stimulation with f-Met-Leu-Phe is three to four fold higher in comparison with neutrophils of normotensive control subjects. Neutrophils from hypertensive patients are less responsive to priming, by non-stimulating doses of the agonist, as compared to control cells, which following this pretreatment augment superoxide anion production up to levels close to those expressed by neutrophils from hypertensive patients. No difference in NADPH oxidase activity, between neutrophils from the two groups of subjects, was observed when the rate of O2- production was evaluated in a reconstructed cell-free system containing the membrane fraction and the cytosolic cofactors. These results are consistent with the hypothesis that differences in the functional organization of the oxidase at the membrane level in neutrophils of hypertensive are responsible for the enhanced O2- production following agonist stimulation.
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PMID:Enhanced activation of the respiratory burst oxidase in neutrophils from hypertensive patients. 253 41

The NADPH:O2 oxidoreductase (NADPH oxidase) of human neutrophils is converted from a dormant to an active state upon stimulation of the cells. We have studied the soluble fraction that is required for NADPH oxidase activation in a cell-free system. Human neutrophils were separated in a membrane-containing and a soluble fraction. The soluble fraction was separated on carboxymethyl (CM) Sepharose in 10 mM 4-morpholino-ethanesulfonic acid buffer of pH 6.8. Reconstitution of the NADPH oxidase activity, measured as O2 consumption, was only found when the membrane fraction was combined with the flowthrough of the CM Sepharose column as well as with a fraction that eluted at 125 mM NaCl. This result indicates that at least two soluble components are necessary for reconstitution of the NADPH oxidase activity: one that does not bind to CM Sepharose and one that does bind. These components were designated soluble oxidase component (SOC) I and SOC II, respectively. Boiling destroyed the activity in both fractions. In the soluble fraction of human lymphocytes and thrombocytes neither SOC I nor SOC II activity was found. SOC II copurified with a 47-kD phosphoprotein, previously found defective in patients with the autosomal form of chronic granulomatous disease (CGD). Inactive soluble fractions of cells from autosomal CGD patients were reconstituted with a SOC II fraction from control cells. The result of this experiment indicates that autosomal CGD patients are normal in SOC I but defective in SOC II.
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PMID:A phosphoprotein of Mr 47,000, defective in autosomal chronic granulomatous disease, copurifies with one of two soluble components required for NADPH:O2 oxidoreductase activity in human neutrophils. 253 48


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