共查询到20条相似文献,搜索用时 31 毫秒
1.
1. The NADPH-oxidizing activity of a 100 000 × g particulate fraction of the postnuclear supernatant obtained from guinea-pig phagocytosing polymorphonuclear leucocytes has been assayed by simultaneous determination of oxygen consumption, NADPH oxidation and O?2 generation at pH 5.5 and 7.0 and with 0.15 mM and 1 mM NADPH.2. The measurements of oxygen consumption and NADPH oxidation gave comparable results. The stoichiometry between the oxygen consumed and the NADPH oxidized was 1 : 1.3. A markedly lower enzymatic activity was observed, under all the experimental conditions used, when the O?2 generation assay was employed as compared to the assays of oxygen uptake and NADPH oxidation.4. The explanation of this difference came from the analysis of the effect of superoxide dismutase and of cytochrome c which removes O?2 formed during the oxidation of NADPH.5. Both superoxide dismutase and cytochrome c inhibited the NADPH-oxidizing reaction at pH 5.5. The inhibition was higher with 1 mM NADPH than with 0.15 mM NADPH.6. Both superoxide dismutase and cytochrome c inhibited the NADPH-oxidizing reaction at pH 7.0 with 1 mM NADPH but less than at pH 5.5 with 1 mM NADPH.7. The effect of superoxide dismutase at pH 7.0 with 0.15 mM NADPH was negligible.8. In all instances the inhibitory effect of cytochrome c was greater than that of superoxide dismutase.9. It was concluded that the NADPH-oxidizing reaction studied here is made up of three components: an enzymatic univalent reduction of O2; an enzymatic, apparently non-univalent, O2 reduction and a non-enzymatic chain reaction.10. These three components are variably and independently affected by the experimental conditions used. For example, the chain reaction is freely operative at pH 5.5 with 1 mM NADPH but is almost absent at pH 7.0 with 0.15 mM NADPH, whereas the univalent reduction of O2 is optimal at pH 7.0 with 1 mM NADPH. 相似文献
2.
The stoichiometry of hydroxylation reactions catalyzed by cytochrome P-450 was studied in a reconstituted enzyme system containing the highly purified cytochrome from phenobarbital-induced rabbit liver microsomes. Hydrogen peroxide was shown to be formed in the reconstituted system in the presence of NADPH and oxygen; the amount of peroxide produced varied with the substrated added. NADPH oxidation, oxygen consumption, and total product formation (sum of hydroxylated compound and hydrogen peroxide) were shown to be equimolar when cyclohexane, benzphetamine, or dimethylaniline served as the substrate. The stoichiometry observed represents the sum of two activities associated with cytochrome P-450. These are (1) hydroxylase activity: NADPH + H+ + O2 + RH → NADP+ + H2O + ROH; and (2) oxidase activity: NADPH + H+ + O2 → NADP+ + H2O2. Benzylamphetamine (desmethylbenzphetamine) acts as a pseudosubstrate in that it stimulates peroxide formation to the same extent as the parent compound (benzphetamine), but does not undergo hydroxylation. Accordingly, when benzylamphetamine alone is added in control experiments to correct for the NADPH and O2 consumption not associated with benzphetamine hydroxylation, the expected 1:1:1 stoichiometry for NADPH oxidation, O2 consumption, and formaldehyde formation in the hydroxylation reaction is observed. 相似文献
3.
It is postulated that the burst of oxygen consumption and H2O2 formation following phagocytosis by polymorphonuclear leukocytes is due to the action of an oxidase located in the plasma membrane. The cyanide-resistant oxygen consumption of resting polymorphonuclear leukocytes was also found to be stimulated by 2,4-dichlorophenol with H2O2 being the sole product formed. NADH and NADPH added to the leukocytes greatly enhanced the oxygen consumption and were oxidized in the process without penetrating the leukocytes. Mn2+ stimulated this oxidase activity. The apparent Km values for added NADH and NADPH were 50 and 40 μm, respectively, with a V of 300 nmol/mg protein/min. A stoichiometry of 1 mol H2O2 formed per mol of NAD(P)H was found. Whilst the oxidase is similar to the oxidase properties of a peroxidase, myeloperoxidase is not responsible for the activity. 相似文献
4.
Timothy L. Macdonald Leo T. Burka S.Tracy Wright F.Peter Guengerich 《Biochemical and biophysical research communications》1982,104(2):620-625
Highly-purified rat liver microsomal cytochrome P-450 converted cyclohexane to cyclohexanol in the presence of iodosobenzene. Oxygen from 18O-iodosobenzene was not incorporated into cyclohexanol but oxygen from H218O was readily incorporated. Cytochrome P-450 catalyzed the facile exchange of oxygen between iodosobenzene and water but neither cytochrome P-420 nor the apoenzyme did. Under these conditions cytochrome P-450 readily incorporated oxygen from 18O2 into cyclohexanol in the presence of NADPH-cytochrome P-450 reductase and NADPH. The results are interpreted in a mechanism in which cytochrome P-450 forms a common hydroxylating species in the presence of iodosobenzene or O2 plus NADPH. 相似文献
5.
Various factor affecting NADPH-oxidation by resting human leucocyte granules (LG) at acid pH, have been investigated.It was found that:
- 1) oxidation of NADPH by LG was increasingly inhibited by increased cyanide concentrations in the medium and was abolished by 4 mM cyanide.
- 2) with or without cyanide in the incubation medium, LG omitted, Mn++, in the presence of NADPH induced superoxide anion (O¯2) production, as evidenced by oxygen consumption and H2O2 production, which were abolished (in the absence of cyanide) by cytochrome C (a potent O¯2 scavenger).
- 3) Both NADPH oxidation in the presence of 2 mM cyanide (cyanide-resistant) and in its absence (cyanide-sensitive) by LG occured only in the presence of Mn++, and both were inhibited by superoxide dismutase.
- 4) Cyanide-resistant NADPH oxidation by LG generated H2O2, was inhibited by H2O2 and was not modified by «active catalase. The ratio of cyanide-resistant NADPH oxidation/O2 uptake was 1 up to 1.25 mM NADPH, and increased above this concentration.
- 5) Cyanide-sensitive NADPH oxidation was inhibited by catalase and increased upon addition of H2O2. The ratio of cyanide-sensitive NADPH oxidation/O2 uptake was 2.
6.
The addition of nitrofurantoin to aerobic incubation mixtures containing rat lung microsomes strongly enhanced the generation of adrenochrome from epinephrine. Adrenochrome formation in this system was blocked by superoxide dismutase, but not by catalase. Hydrogen peroxide production was also strongly enhanced by nitrofurantoin in these preparations; superoxide dismutase did not significantly alter the amount of H2O2 measured, but no H2O2 was detected in incubation mixtures in the presence of catalase. Nitrofurantoin enhanced the oxidation of NADPH in lung microsomal suspensions under aerobic conditions; the enhancement was unaffected by catalase but was partially prevented by superoxide dismutase. Neither adrenochrome formation nor H2O2 production were enhanced by nitrofurantoin under anaerobic (N2) conditions, but NADPH oxidation in the presence of nitrofurantoin was greater under anaerobic conditions than under aerobic conditions. These results are consistent with the view that the redox cycling of nitrofurantoin in lung microsomes in the presence of oxygen results in the consumption of NADPH and the production of activated oxygen species, emphasizing some metabolic similarities with the lung-toxic herbicide, paraquat. 相似文献
7.
K Sugioka H Nakano T Noguchi J Tsuchiya M Nakano 《Biochemical and biophysical research communications》1981,100(3):1251-1258
The system, which contains NADPH, purified cytochrome P-450 reductase, and adriamycin, produces H2O2 and in appreciable amounts with oxygen consumption and NADPH oxidation under aerobic conditions. Such an adriamycin-induced NADPH oxidation system, however, does not cause the decomposition of unsaturated fatty acids in microsomal phospholipid micelles, suggesting no direct participation of the active oxygen species and semiquinone radicals of adriamycin in lipid peroxidation. Adriamycin produces a co-ordination complex with Fe3+ and ADP, which, but no Fe3+-ADP complex, could be reduced by NADPH-cytochrome P-450 reductase at the expence of NADPH. The decomposition of unsaturated fatty acids in phospholipid micelles is achieved by the Fe3+-ADP-adriamycin complex and strikingly enhanced by enzymatically reduced iron-ADP-adriamycin complex. 相似文献
8.
Nitroalkene derivative of oleic acid (OA-NO2), due to its ability to mediate revisable Michael addition, has been demonstrated to have various biological properties and become a therapeutic agent in various diseases. Though its antioxidant properties have been reported in different models of acute kidney injury (AKI), the mechanism by which OA-NO2 attenuates intracellular oxidative stress is not well investigated. Here, we elucidated the anti-oxidative mechanism of OA-NO2 in an in vitro model of renal ischemia/reperfusion (I/R) injury. Human tubular epithelial cells were subjected to oxygen and glucose deprivation/re-oxygenation (OGD/R) injury. Pretreatment with OA-NO2 (1.25?μM, 45?min) attenuated OGD/R triggered reactive oxygen species (ROS) generation and subsequent mitochondrial membrane potential disruption. This action was mediated via up-regulating endogenous antioxidant defense components including superoxide dismutase (SOD1), heme oxygenase 1 (HO-1), and γ-glutamyl cysteine ligase modulatory subunits (GCLM). Moreover, subcellular fractionation analyses demonstrated that OA-NO2 promoted nuclear translocation of nuclear factor-E2- related factor-2 (Nrf2) and Nrf2 siRNA partially abrogated these protective effects. In addition, OA-NO2 inhibited NADPH oxidase activation and NADPH oxidase 4 (NOX4), NADPH oxidase 2 (NOX2) and p22phox up-regulation after OGD/R injury, which was not relevant to Nrf2. These results contribute to clarify that the mechanism of OA-NO2 reno-protection involves both inhibition of NADPH oxidase activity and induction of SOD1, Nrf2-dependent HO-1, and GCLM. 相似文献
9.
《Free radical research》2013,47(4-6):271-277
When NADPH-cytochrome P-450 reductase isolated from rat liver microsomes was aerobically incubated with bleomycin, FeCl3, NADPH and DNA parallel NADPH and oxygen were consumed and malondialdehyde was formed. A similar parallelism of NADPH- and oxygen-consumption and malondialdehyde formation was observed when ceil nuclei isolated from rat liver were incubated under the same conditions. The formation of malondialdehyde which was identified by HPLC and which was most likely released from oxidative cleavage of deoxyribose of nuclear DNA required oxygen, bleomycin, FeCl3 and NADPH. This indicates that a nuclear NADPH-enzyme, presumably NADPH-cytochrome P-450 reductase, is able to redox cycle a bleomycin-iron-complex which in the reduced form can activate oxygen to a DNA-damaging reactive species. The data suggest that the activity of this enzyme in the cell nucleus could play an important role in the cytotoxicity of bleomycin in tumor cells. 相似文献
10.
Ye J Ding M Leonard SS Robinson VA Millecchia L Zhang X Castranova V Vallyathan V Shi X 《Molecular and cellular biochemistry》1999,202(1-2):9-17
Apoptosis is a physiological mechanism for the control of DNA integrity in mammalian cells. Vanadium induces both DNA damage and apoptosis. It is suggested that vanadium-induced apoptosis serves to eliminate DNA-damaged cells. This study is designed to clarify a role of reactive oxygen species in the mechanism of apoptosis induced by vanadium. We established apoptosis model with murine epidermal JB6 P+ cells in the response to vanadium stimulation. Apoptosis was detected by a cell death ELISA assay and morphological analysis. The result shows that apoptosis induced by vanadate is dose-dependent, reaching its saturation level at a concentration of 100 M vanadate. Vanadyl (IV) can also induce apoptosis albeit with lesser potency. A role of reactive oxygen species was analyzed by multiple reagents including specific scavengers of different reactive oxygen species. The result shows that vanadate-induced apoptosis is enhanced by NADPH, superoxide dismutase and sodium formate, but was inhibited by catalase and deferoxamine. Cells exposed to vanadium consume more molecular oxygen and at the same time, produce more H2O2 as measured by the change in fluorescence of scopoletin in the presence of horseradish peroxidase. This change in oxygen consumption and H2O2 production is enhanced by NADPH. Taken together, these results show that vanadate induces apoptosis in epidermal cells and H2O2 induced by vanadate plays a major role in this process. 相似文献
11.
Fujimori N Oono T Igarashi H Ito T Nakamura T Uchida M Coy DH Jensen RT Takayanagi R 《Peptides》2011,32(10):2067-2076
Vasoactive intestinal peptide (VIP) attenuates experimental acute pancreatitis (AP) by inhibition of cytokine production from inflammatory cells. It has been suggested that reactive oxygen species (ROS) as well as cytokines play pivotal roles in the early pathophysiology of AP. This study aimed to clarify the effect of VIP on the oxidative condition in pancreas, especially pancreatic acinar cells (acini). Hydrogen peroxide (H2O2)-induced intracellular ROS, assessed with CM-H2DCFDA, increased time- and dose-dependently in acini isolated from rats. Cell viability due to ROS-induced cellular damage, evaluated by MTS assay, was decreased with ≥100 μmol/L H2O2. VIP significantly inhibited ROS production from acini and increased cell viability in a dose-dependent manner. Expression of antioxidants including catalase, glutathione reductase, superoxide dismutase (SOD) 1 and glutathione peroxidase was not altered by VIP except for SOD2. Furthermore, Nox1 and Nox2, major components of NADPH oxidase, were expressed in pancreatic acini, and significantly increased after H2O2 treatment. Also, NADPH oxidase activity was provoked by H2O2. VIP decreased NADPH oxidase activity, which was abolished by PKA inhibitor H89. These results suggested that VIP affected the mechanism of ROS production including NADPH oxidase through induction of a cAMP/PKA pathway. In conclusion, VIP reduces oxidative stress in acini through the inhibition of NADPH oxidase. These results combined with findings of our previous study suggest that VIP exerts its protective effect in pancreatic damage, not only through an inhibition of cytokine production, but also through a reduction of the injury caused by oxidative stress. 相似文献
12.
Jinyoung Hur Mi Jung Kim Young-Wuk Cho 《Biochemical and biophysical research communications》2010,391(3):1526-59
Although glial cells play a major role in the pathogenesis of many neurological diseases by exacerbating neuronal and non-neuronal cell death, the mechanisms involved are unclear. We examined the effects of microglia-(MCM) or astrocyte-(ACM) conditioned media obtained by chemical ischemia on the neuronal injury in SH-SY5Y cells. Chemical ischemia was induced by the treatment with NaN3 and 2-deoxy-d-glucose for 2 h. MCM-treated SH-SY5Y cells showed reduced the viability, increased caspase-3 activity, decreased Bcl-2/Bax ratio, and increased cytochrome c release, increased inflammatory cytokines, and increased reactive oxygen species (ROS) generation. MCM also increased gp91phox nicotinamide adenine dinucleotide phosphate (NADPH) oxidase, which was inhibited by NADPH oxidase inhibitor, apocynin, and gp91phox siRNA. However, ACM did not show any significant changes. The results suggest that microglia activated by ischemic insult may increase reactive oxygen species generation via activation of gp91phox NADPH oxidase, resulting in neuronal injury. 相似文献
13.
14.
《Journal of Fermentation Technology》1987,65(4):483-487
A lignin-degrading basidiomycete, Phanerochaete chrysosporium, produces an extracellular peroxidase which in turn produces H2O2 by catalyzing the oxidation of NADH and NADPH. The high enzyme activity was observed in the culture grown under nutrient nitrogen limitation (low-N) and high oxygen tension (high-O2). The enzyme activity was absent in non-ligninolytic agitated culture and in the cultures of non-ligninolytic mutant strains of this organism. The culture method using polyurethane foam cubes as a support for the growing mycelia showed the beneficial effect of producing a large amount of the enzyme. The enzyme is capable of catalyzing the oxidation of NADH and NADPH in the absence of added H2O2, and its activity was inhibited strongly by catalase and superoxide dismutase. It is suggested that this peroxidase participates in the ligninolytic system of Phanerochaete chrysosporium as a donor of H2O2, which is required for the lignin-peroxidase reaction, by oxidizing extracellular NADH and NADPH. 相似文献
15.
Theodore G. Gabig Bruce A. Lefker 《Biochemical and biophysical research communications》1984,118(2):430-436
The resolved flavoprotein and cytochrome b559 components of the NADPH dependent generating oxidase from human neutrophils were the subject of further study. The resolved flavoprotein, depleted of cytochrome b559, was reduced by NADPH under anaerobic conditions and reoxidized by oxygen. NADPH dependent generation by the resolved flavoprotein fraction was not detectable, however it was competent in the transfer of electrons from NADPH to artificial electron acceptors. The resolved cytochrome b559, depleted of flavoprotein, demonstrated no measureable NADPH dependent generating activity and was not reduced by NADPH under anaerobic conditions. The dithionite reduced form of the resolved cytochrome b559 was rapidly oxidized by oxygen, as was the cytochrome b559 in the intact oxidase. 相似文献
16.
Kim JY Yu SJ Oh HJ Lee JY Kim Y Sohn J 《Apoptosis : an international journal on programmed cell death》2011,16(4):347-358
Panaxydol, a polyacetylenic compound derived from Panax ginseng roots, has been shown to inhibit the growth of cancer cells. In this study, we demonstrated that panaxydol induced apoptosis
preferentially in transformed cells with a minimal effect on non-transformed cells. Furthermore, panaxydol was shown to induce
apoptosis through an increase in intracellular Ca2+ concentration ([Ca2+]i), activation of JNK and p38 MAPK, and generation of reactive oxygen species (ROS) initially by NADPH oxidase and then by
mitochondria. Panaxydol-induced apoptosis was caspase-dependent and occurred through a mitochondrial pathway. ROS generation
by NADPH oxidase was critical for panaxydol-induced apoptosis. Mitochondrial ROS production was also required, however, it
appeared to be secondary to the ROS generation by NADPH oxidase. Activation of NADPH oxidase was demonstrated by the membrane
translocation of regulatory p47phox and p67phox subunits and shown to be necessary for ROS generation by panaxydol treatment. Panaxydol triggered a rapid and sustained increase
of [Ca2+]i, which resulted in activation of JNK and p38 MAPK. JNK and p38 MAPK play a key role in activation of NADPH oxidase, since
inhibition of their expression or activity abrogated membrane translocation of p47phox and p67phox subunits and ROS generation. In summary, these data indicate that panaxydol induces apoptosis preferentially in cancer cells,
and the signaling mechanisms involve a [Ca2+]i increase, JNK and p38 MAPK activation, and ROS generation through NADPH oxidase and mitochondria. 相似文献
17.
Addition of nifurtimox (a nitrofuran derivative used for the treatment of Chagas' disease) to rat liver microsomes produced an increase of (a) electron flow from NADPH to molecular oxygen, (b) generation of both superoxide anion radical (O2?) and hydrogen peroxide, and (c) lipid peroxidation. The nifurtimox-stimulated NADPH oxidation was greatly inhibited by NADP+ and p-chloromercuribenzoate, and to a lesser extent by SKF-525-A and metyrapone. These inhibitions reveal the function of both the NADPH-cytochrome P-450 (c) reductase and cytochrome P-450 in nifurtimox reduction. Superoxide dismutase, catalase (in the presence of superoxide dismutase), and hydroxyl radical scavengers (mannitol, 5,5-dimethyl-1-pyrroline-1-oxide) inhibited the nifurtimox-stimulated NADPH oxidation, in accordance with the additional operation of a reaction chain including the hydroxyl radical. Further evidence supporting the role of superoxide anion and hydroxyl radicals in the nifurtimox-induced NADPH oxidation resulted from the effect of specific inhibitors on NADPH oxidation by O2? (generated by the xanthine oxidase reaction) and by OH. (generated by an iron chelate or the Fenton reaction). Production of O2? by rat kidney, testes and brain microsomes was significantly stimulated by nifurtimox in the presence of NADPH. It is postulated that enhanced formation of free radicals is the basis for nifurtimox toxicity in mammals, in good agreement with the postulated mechanism of the trypanocide effect of nifurtimox on Trypanosoma cruzi. 相似文献
18.
NADPH reduced rabbit liver microsomal enzymes catalyzed anaerobic dehalogenation of halothane (2-bromo-2-chloro-1,1,1-trifluoroethane) to produce CF2CHCl and CF3CH2Cl. Anaerobic dehalogenation was optimal at pH7.4 and was blocked by either oxygen or carbon monoxide. The degree of inhibition of anaerobic dehalogenation by carbon monoxide was closely correlated to the proportion of carbon monoxide complex of cytochrome P450. Anaerobic dehalogenation was enhanced by pretreatment of the animals with phenobarbital but not with methylcholanthrene. 相似文献
19.
O Augusto E J Bechara D L Sanioto G Cilento 《Archives of biochemistry and biophysics》1973,158(1):359-364
Catechol and catecholamines have been assayed upon the microsomal NADPH and NADH oxidase activities. Epinephrine shows a catalytic effect on the NADPH oxidation characterized by a small lag. The two to threefold increase in rate can be suppressed by Superoxide dismutase if the enzyme is added before the reaction begins. The catalytic effect is ascribed to a quinone formed by two electron oxidation of epinephrine by the Superoxide ion. The quinone, which is not catalytically active in the NADH chain, appears to mediate electrons between the NADPH-cytochrome c reductase and oxygen. The four electron oxidation product adrenochrome is also active upon the NADPH chain but inactive upon the NADH chain.Epinephrine did not change the menadione-stimulated NADPH oxidase activity. Presumably, during this and the NADH oxidase activities, two electrons are simultaneously transferred to the oxygen molecule.Catechol and catecholamines doubled the rate of autoxidation of NADH in the presence of catalytic amounts of NADH-cytochrome b5 reductase and cytochrome b5, a result which suggests Superoxide ion formation in the autoxidation of the cytochrome.Epinephrine does not act upon the desaturation of endogenous substrate or upon endogenous lipid peroxidation. 相似文献
20.
Maria Tomasz 《Chemico-biological interactions》1976,13(1):89-97
Reduction of mitomycin C by NaBH4 or by NADPH in the presence of a cell extract followed by exposure to air results in the generation of H2O2. This phenomenon occurs not only with free mitomycin but also with mitomycin irreversibly bound to DNA. In view of these findings, the antibiotic activity of mitomycin was tested in two bacterial systems: a facultative aerobic bacterium grown in the presence or absence of oxygen and an obligate anaerobic bacterium. No oxygen effect could be demonstrated in either case in the growth-inhibitory and bactericidal activity of the drug. Nevertheless, the H2O2 generating capacity of mitomycin-DNA complexes inside the nucleus may play a role in the drug-induced biological damage to the genetic material of cells. 相似文献