Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:1.6.99.5 (NADH dehydrogenase)
 2,135 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Modeling of ischemic phenomena in vitro has been hindered by the inability to create specific alterations in the variables of interest over a defined time-frame. In particular, changes in the adenine nucleotide pool have been quite difficult to mimic because of the putative low metabolic rate in culture and the long times necessary to achieve even partial chemical energy depletion. Here we present evidence for a rapid method of producing a profound chemical energy depletion with the combination of a NADH dehydrogenase inhibitor (amytal) and a mitochondrial proton ionophore (CCCP). Treatment with our protocol in enriched spinal cultures results in a 40% decrease in ATP within 2 min and a fall to one-third of control values by 15 min. The overall pool size of the total adenine nucleotides is decreased 46% by 15 min and does not completely recover after 5 min of reenergization. The ATP/ADP ratio declines to one-third of control values during deenergization and returns to control values after 5 min in control buffer. Such a loss of the total adenylate pool closely mimics that seen in vivo during ischemia and provides an in vitro model system in which the effects of the combination of this means of cellular injury with others (e.g., excitotoxins) may be examined.
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PMID:Energy depletion in culture. Adenine nucleotides are altered as in vivo. 177 32

Ischemia and reperfusion causes severe mitochondrial damage, including swelling and deposits of hydroxyapatite crystals in the mitochondrial matrix. These crystals are indicative of a massive influx of Ca2+ into the mitochondrial matrix occurring during reoxygenation. We have observed that mitochondria isolated from rat hearts after 90 minutes of anoxia followed by reoxygenation, show a specific inhibition in the electron transport chain between NADH dehydrogenase and ubiquinone in addition to becoming uncoupled (unable to generate ATP). This inhibition is associated with an increased H2O2 formation at the NADH dehydrogenase level in the presence of NADH dependent substrates. Control rat mitochondria exposed for 15 minutes to high Ca2+ (200 nmol/mg protein) also become uncoupled and electron transport inhibited between NADH dehydrogenase and ubiquinone, a lesion similar to that observed in post-ischemic mitochondria. This Ca(2+)-dependent effect is time dependent and may be partially prevented by albumin, suggesting that it may be due to phospholipase A2 activation, releasing fatty acids, leading to both inhibition of electron transport and uncoupling. Addition of arachidonic or linoleic acids to control rat heart mitochondria, inhibits electron transport between Complex I and III. These results are consistent with the following hypothesis: during ischemia, the intracellular energy content drops severely, affecting the cytoplasic concentration of ions such as Na+ and Ca2+. Upon reoxygenation, the mitochondrion is the only organelle capable of eliminating the excess cytoplasmic Ca2+ through an electrogenic process requiring oxygen (the low ATP concentration makes other ATP-dependent Ca2+ transport systems non-operational).(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:Mitochondrial generation of oxygen radicals during reoxygenation of ischemic tissues. 206 Aug 40

The hypothesis that mitochondria damaged during complete cerebral ischemia generate increased amounts of superoxide anion radical and hydrogen peroxide (H2O2) upon postischemic reoxygenation has been tested. In rat brain mitochondria, succinate supported H2O2 generation, whereas NADH-linked substrates, malate plus glutamate, did so only in the presence of respiratory chain inhibitors. Succinate-supported H2O2 generation was diminished by rotenone and the uncoupler carbonyl cyanide m-chlorphenylhydrazone and enhanced by antimycin A and increased oxygen tensions. When maximally reduced, the NADH dehydrogenase and the ubiquinone-cytochrome b regions of the electron transport chain are sources of H2O2. These studies suggest that a significant portion of H2O2 generation in brain mitochondria proceeds via the transfer of reducing equivalents from ubiquinone to the NADH dehydrogenase portion of the electron transport chain. Succinate-supported H2O2 generation by mitochondria isolated from rat brain exposed to 15 min of postdecapitative ischemia was 90% lower than that of control preparations. The effect of varying oxygen tensions on H2O2 generation by postischemic mitochondrial preparations was negligible compared with the increased H2O2 generation measured in control preparations. Comparison of the effects of respiratory chain inhibitors and oxygen tension on succinate-supported H2O2 generation suggests that the ability for reversed electron transfer is impaired during ischemia. These data do not support the hypothesis that mitochondrial free radical generation increases during postischemic reoxygenation.
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PMID:Generation of hydrogen peroxide by brain mitochondria: the effect of reoxygenation following postdecapitative ischemia. 291 86

Complex I (NADH-ubiquinone reductase) is a complex system located in the inner mitochondrial membrane and has the ability to catalyse several different enzymatic reactions concerned in electron transport. It is known to be one of the first components of the respiratory chain to be damaged by ischemia. Our results, using autolysis in the rat heart as experimental model, indicate that the NADH dehydrogenase system was impaired relatively early during ischemia while transhydrogenation and NADPH dehydrogenation appeared to be relatively resistant.
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PMID:Changes in NADH-ubiquinone reductase (complex I) with autolysis in the rat heart as experimental model. 309 11

The status of glutathione (GSH) and protein thiol homeostasis was examined in rat brain regions during reperfusion after moderate and severe cerebral ischemia. GSH levels were decreased in brain regions during reperfusion for 1 hr after moderate or severe ischemia for 0.5 hr. Maximal loss of GSH (50-66%) was observed in the striatum and hippocampus. The GSH lost from the brain regions was essentially recovered as protein-glutathione mixed disulfide (PrSSG) with concomitant loss of protein thiols (PrSH). The activities of enzymes such as Na+K+ ATPase, NADH dehydrogenase and glutathione reductase were also inhibited but were restored after incubation of the brain homogenate with dithiothreitol. The depletion of GSH was also accompanied by an increase in the levels of malondialdehyde and reactive oxygen species. The total GSH recovered as sum of GSH and PrSSG was significantly higher than the sham-operated controls in the hippocampus and striatum after 1 hr of reperfusion, after moderate ischemia for 0.5 hr, and at the end of 24 hr of reperfusion the GSH-protein thiol homeostasis was restored. In contrast after 1 hr of reperfusion after severe ischemia, the GSH recovered as sum of GSH and PrSSG was not significantly different from sham-operated controls and at the end of 24 hr, 7 of 9 animals died. The recuperation of the brain from oxidative stress during reperfusion after moderate ischemia was thus preceded by increased recovery of total GSH essentially in the form of PrSSG. Thus, rapid restoration of thiol homeostasis in the brain during reperfusion may help the brain recover from reperfusion injury.
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PMID:Glutathione and protein thiol homeostasis in brain during reperfusion after cerebral ischemia. 756 84

Electron transport and production of O2-/H2O2 by the NADH dehydrogenase flavin-semiquinone (FMNH.) and ubisemiquinone (UQH.) were studied in a model of in vivo ischemia-reperfusion in rat kidney. H2O2 production rates were assessed in isolated mitochondria using either succinate, with and without antimycin, or malate-glutamate, with and without rotenone. Respiratory activities of isolated mitochondria and activity of NADH- and succinate-cytochrome c reductase and of NADH- and succinate-dehydrogenase in submitochondrial particles were measured to evaluate the electron flux throughout respiratory carriers. The mitochondrial H2O2 production rate was approximately 1.5- and 4-times increased in ischemic and ischemic-reperfused kidneys, respectively. Ischemia caused a marked decrease in the electron transport throughout the NADH-UQ segment with no significant changes either in the NADH dehydrogenase activity or in the electron flux trough the succinate-cytochrome oxidase segment. Reperfusion did not further affect the NADH-ubiquinone segment but markedly inhibited the succinate-supported oxygen consumption, succinate-cytochrome c reductase and succinate dehydrogenase activity. Our results show a redistribution of the electron flux with an increased rate of superoxide anion/hydrogen peroxide production at NADH dehydrogenase in mitochondria subjected to ischemia only. After 10 min reperfusion an impairment of the electron flow at succinate-cytochrome c segment is established and hydrogen peroxide production by UQH. increases up to maximal values becoming the major source of superoxide anion/hydrogen peroxide.
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PMID:Mitochondrial sites of hydrogen peroxide production in reperfused rat kidney cortex. 772 10

Previous in vitro studies have shown that isolated mitochondria can generate oxygen radicals. However, whether a similar phenomenon can also occur in intact organs is unknown. In the present study, we tested the hypothesis that resumption of mitochondrial respiration upon reperfusion might be a mechanism of oxygen radical formation in postischemic hearts, and that treatment with inhibitors of mitochondrial respiration might prevent this phenomenon. Three groups of Langendorff-perfused rabbit hearts were subjected to 30 min of global ischemia at 37 degrees C, followed by reflow. Throughout ischemia and early reperfusion the hearts received, respectively: (a) 5 mM KCl (controls), (b) 5 mM sodium amobarbital (Amytal, which blocks mitochondrial respiration at Site I, at the level of NADH dehydrogenase), and (c) 5 mM potassium cyanide (to block mitochondrial respiration distally, at the level of cytochrome c oxidase). The hearts were then processed to directly evaluate oxygen radical generation by electron paramagnetic resonance spectroscopy, or to measure oxygen radical-induced membrane lipid peroxidation by malonyl dialdehyde (MDA) content of subcellular fractions. Severity of ischemia, as assessed by 31P-nuclear magnetic resonance measurements of cardiac ATP, phosphocreatine, and pH, was similar in all groups. Oxygen-centered free radical concentration averaged 3.84 +/- 0.54 microM in reperfused control hearts, and it was significantly reduced by Amytal treatment (1.98 +/- 0.26; p < 0.05), but not by KCN (2.58 +/- 0.96 microM; p = not significant (NS)), consistent with oxygen radicals being formed in the mitochondrial respiratory chain at Site I. Membrane lipid peroxidation of reperfused hearts was also reduced by treatment with Amytal, but not with KCN. MDA content of the mitochondrial fraction averaged 0.75 +/- 0.06 nM/mg protein in controls, 0.72 +/- 0.06 in KCN-treated hearts, and 0.54 +/- 0.05 in Amytal-treated hearts (p < 0.05 versus both groups). Similarly, MDA content of lysosomal membrane fraction was 0.64 +/- 0.09 nM/mg protein in controls, 0.79 +/- 0.15 in KCN-treated hearts, and 0.43 +/- 0.06 in Amytal-treated hearts (p < 0.05 versus both groups). Since the effects of Amytal are known to be reversible, in a second series of experiments we investigated whether transient mitochondrial inhibition during the initial 10 min of reperfusion was also associated with beneficial effects on subsequent recovery of cardiac function after wash-out of the drug. At the end of the experiment, recovery of left ventricular end-diastolic and of developed pressure was significantly greater in those hearts that had been treated with Amytal during ischemia and early reflow, as compared to untreated hearts.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Evidence that mitochondrial respiration is a source of potentially toxic oxygen free radicals in intact rabbit hearts subjected to ischemia and reflow. 839 7

The mitochondria harvested at the end of perfusion of control hearts and assayed for respiratory activity had a better function after ischemia and reperfusion following trimetazidine injection when glutamate was used as substrate. The protective effect of trimetazidine was enhanced when the mitochondria were isolated from hypertrophied perfused rat hearts. In fact the drug improved both the RCI and QO2 parameters with glutamate or succinate as substrates and raised the glutamate-induced QO2 value of mitochondria extracted from the hypertrophied heart perfused in aerobic conditions. In the aerobically perfused heart trimetazidine did not change either the levels of tissue malondialdehyde and lipofuscin, or the rate of mitochondrial O.2 generation while it reduced the O.2 formation and malondialdehyde content in the hypertrophied heart. After ischemia and reperfusion, the drug reproduced these protective effects in the hypertrophied hearts and reduced the level of tissue malondialdehyde in control hearts. The protective effect of trimetazidine against MDA formation was dose-dependent, being more evident at a higher dose (10 mumol/l). Preincubation of rat heart mitochondria with 0.1-10 mumol/l trimetazidine did not affect NADH oxidase, NADH dehydrogenase and NADH-cytochrome c reductase, succinate oxidase and cytochrome c oxidase activities. These results indicate that trimetazidine injected into isolated rat hearts protects against the damage induced on cardiac energetics and oxidative injuries by moderate ischemia and reperfusion stress, particularly in monocrotaline-induced hypertrophy in the rat heart. We suggest that trimetazidine reduces the formation of oxidative damage by preserving cardiac mitochondrial function.
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PMID:Effect of trimetazidine on mitochondrial function and oxidative damage during reperfusion of ischemic hypertrophied rat myocardium. 851 81

The mitochondrial respiratory chain is a major source of reactive oxygen species (ROS) under pathological conditions including myocardial ischemia and reperfusion. Limitation of electron transport by the inhibitor rotenone immediately before ischemia decreases the production of ROS in cardiac myocytes and reduces damage to mitochondria. We asked if ROS generation by intact mitochondria during the oxidation of complex I substrates (glutamate, pyruvate/malate) occurred from complex I or III. ROS production by mitochondria of Sprague-Dawley rat hearts and corresponding submitochondrial particles was studied. ROS were measured as H2O2 using the amplex red assay. In mitochondria oxidizing complex I substrates, rotenone inhibition did not increase H2O2. Oxidation of complex I or II substrates in the presence of antimycin A markedly increased H2O2. Rotenone prevented antimycin A-induced H2O2 production in mitochondria with complex I substrates but not with complex II substrates. Catalase scavenged H2O2. In contrast to intact mitochondria, blockade of complex I with rotenone markedly increased H2O2 production from submitochondrial particles oxidizing the complex I substrate NADH. ROS are produced from complex I by the NADH dehydrogenase located in the matrix side of the inner membrane and are dissipated in mitochondria by matrix antioxidant defense. However, in submitochondrial particles devoid of antioxidant defense ROS from complex I are available for detection. In mitochondria, complex III is the principal site for ROS generation during the oxidation of complex I substrates, and rotenone protects by limiting electron flow into complex III.
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PMID:Production of reactive oxygen species by mitochondria: central role of complex III. 1284 17

Ischemic preconditioning confers cardiac protection during subsequent ischemia-reperfusion, in which protein kinase C (PKC) is believed to play an essential role, but controversial data exist concerning the PKC-delta isoform. In an accompanying study (26), we described metabolic changes in PKC-delta knockout mice. We now wanted to explore their effect on early preconditioning. Both PKC-delta(-/-) and PKC-delta(+/+) mice underwent three cycles of 5-min left descending artery occlusion/5-min reperfusion, followed by 30-min occlusion and 2-h reperfusion. Unexpectedly, preconditioning exaggerated ischemia-reperfusion injury in PKC-delta(-/-) mice. Whereas ischemic preconditioning increased superoxide anion production in PKC-delta(+/+) hearts, no increase in reactive oxygen species was observed in PKC-delta(-/-) hearts. Proteomic analysis of preconditioned PKC-delta(+/+) hearts revealed profound changes in enzymes related to energy metabolism, e.g., NADH dehydrogenase and ATP synthase, with partial fragmentation of these mitochondrial enzymes and of the E(2) component of the pyruvate dehydrogenase complex. Interestingly, fragmentation of mitochondrial enzymes was not observed in PKC-delta(-/-) hearts. High-resolution NMR analysis of cardiac metabolites demonstrated a similar rise of phosphocreatine in PKC-delta(+/+) and PKC-delta(-/-) hearts, but the preconditioning-induced increase in phosphocholine, alanine, carnitine, and glycine was restricted to PKC-delta(+/+) hearts, whereas lactate concentrations were higher in PKC-delta(-/-) hearts. Taken together, our results suggest that reactive oxygen species generated during ischemic preconditioning might alter mitochondrial metabolism by oxidizing key mitochondrial enzymes and that metabolic adaptation to preconditioning is impaired in PKC-delta(-/-) hearts.
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PMID:Ischemic preconditioning exaggerates cardiac damage in PKC-delta null mice. 1527 9


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