oligomycins and Reperfusion-Injury

Ischemia-reperfusion (I/R) injury causes present challenges in the field of graft transplantation which is also a major contributor to early graft dysfunction or failure after organ transplantation. The study focuses on the effects of prolonged cold-ischemia (CI) on the autophagic activity in the graft lung in a rat orthotopic lung transplantation model.. Donor lungs were preserved under CI conditions for different periods. An orthotopic lung transplantation model was developed, and the lung tissues from donor lungs subjected to CI preservation and reperfusion were harvested. We evaluated the effects of different CI periods on autophagy, reactive oxygen species (ROS) and glucose consumption. Additionally, the mechanism by which prolonged CI affected autophagy was investigated through determination of the molecules related to the mTOR pathway after treatment with 3-Methyladenine (3-MA), rapamycin and an adenosine triphosphate (ATP) synthase inhibitor oligomycin (OM).. Prolonged CI led to increased activities of key glycolytic enzymes, glucose consumption and lactic acid production. Autophagy, ROS and glucose consumption were induced in the graft lung after I/R, which reached peak levels after 6 h and was gradually decreased. Most importantly, the perfusion treatment of 3-MA or OM decreased ROS level and autophagy, but increased the extent of mTOR phosphorylation, while the perfusion treatment of rapamycin induced ROS and autophagy.. Taken together, autophagy mediated by a prolonged CI preservation affects the glucose consumption and ROS production in the graft lung via the mTOR signaling pathway.

Topics: Adenine; Animals; Autophagy; Cold Ischemia; Glycolysis; Lung; Lung Transplantation; Lysosomal Membrane Proteins; Male; Mitochondria; Oligomycins; Organ Preservation; Oxidative Phosphorylation; Perfusion; Rats, Sprague-Dawley; Reactive Oxygen Species; Reperfusion Injury; TOR Serine-Threonine Kinases

We investigated the effects of oligomycin, an F1Fo-ATPase inhibitor, on ischemic acute kidney injury in male and female rats. Ischemic acute kidney injury was induced by clamping the left renal artery and vein for 45 or 60 min followed by reperfusion, 2 weeks after contralateral nephrectomy. Renal dysfunction and histological renal damage were observed 1 day after reperfusion in both male and female rats, although these renal injuries were more marked in male rats than in female rats. Intravenous bolus injection of oligomycin (0.5 mg/kg) 5 min before ischemia markedly attenuated the ischemia/reperfusion-induced renal injury in male rats. However, oligomycin did not show the protective effect in female rats subjected to ischemia/reperfusion-induced renal injury. Pre-ischemic treatment with oligomycin suppressed partly but significantly ischemia-induced renal ATP depletion only in male rats. These results indicate that oligomycin prevents the onset of ischemic acute kidney injury in male but not in female rats, and the effect is accompanied by suppression of the ATP depletion only in the male rat kidney during ischemia, thereby suggesting that the ATP hydrolysis pathway by mitochondrial F1Fo-ATPase induces a sex difference in ischemic acute kidney injury.

Topics: Acute Kidney Injury; Adenosine Triphosphate; Animals; Enzyme Inhibitors; Female; Hydrolysis; Injections, Intravenous; Kidney; Male; Mitochondria; Oligomycins; Proton-Translocating ATPases; Rats; Rats, Sprague-Dawley; Reperfusion Injury; Sex Characteristics

We describe a unique mitochondrial oscillator that depends on oxidative phosphorylation, reactive oxygen species (ROS), and mitochondrial inner membrane ion channels. Cell-wide synchronized oscillations in mitochondrial membrane potential (Delta Psi(m)), NADH, and ROS production have been recently described in isolated cardiomyocytes, and we have hypothesized that the balance between superoxide anion efflux through inner membrane anion channels and the intracellular ROS scavenging capacity play a key role in the oscillatory mechanism. Here, we formally test the hypothesis using a computational model of mitochondrial energetics and Ca(2+) handling including mitochondrial ROS production, cytoplasmic ROS scavenging, and ROS activation of inner membrane anion flux. The mathematical model reproduces the period and phase of the observed oscillations in Delta Psi(m), NADH, and ROS. Moreover, we experimentally verify model predictions that the period of the oscillator can be modulated by altering the concentration of ROS scavengers or the rate of oxidative phosphorylation, and that the redox state of the glutathione pool oscillates. In addition to its role in cellular dysfunction during metabolic stress, the period of the oscillator can be shown to span a wide range, from milliseconds to hours, suggesting that it may also be a mechanism for physiological timekeeping and/or redox signaling.

Topics: Animals; Calcium; Cytosol; Glutathione; Glutathione Peroxidase; Glutathione Reductase; Guinea Pigs; Hydrogen Peroxide; Image Processing, Computer-Assisted; Kinetics; Lasers; Membrane Potentials; Microscopy, Fluorescence; Mitochondria; Models, Biological; Models, Theoretical; Myocytes, Cardiac; Oligomycins; Oscillometry; Oxygen; Photons; Reactive Oxygen Species; Reperfusion Injury; Superoxide Dismutase

Topics: Adenosine Diphosphate; Animals; In Vitro Techniques; Ischemia; Kidney; Kinetics; Mitochondria; Oligomycins; Oxidative Phosphorylation; Oxygen Consumption; Rabbits; Reperfusion Injury; Time Factors; Uncoupling Agents

Toxic oxygen species have been implicated as important mediators of injury after reperfusion of an ischemic organ. The aim of this study was to determine if prior metabolic inhibition, such as that which occurs during ischemia, potentiates oxidant injury in vitro. Bovine pulmonary artery endothelial cells were metabolically inhibited for various periods of time with or without the mitochondrial inhibitor oligomycin (650 nM). The cells were rescued from metabolic inhibition by a wash step and subsequent addition of 5.5 mM glucose. At the same time that metabolic inhibition was relieved the cells were subjected to doses of H2O2 ranging from 0 to 100 microM. ATP levels were monitored over a 2-hr time course after rescue from metabolic inhibition by the luciferin-luciferase assay. Cell viability at 2 hr after relief of metabolic inhibition was assessed by trypan blue exclusion. Intracellular pH during metabolic inhibition was determined with the fluorescent dye 2',7'-bis-(2-carboxyethyl)-5(and-6) carboxyfluorescein tetraacetomethoxymethyl ester. H2O2 consumption, a measure of H2O2 scavenging capability, was determined by a fluorescent assay. The viability and ATP levels of cells not subjected to metabolic inhibition were unaffected by these low concentrations of H2O2. Cells metabolically inhibited with glucose depletion and oligomycin were exquisitely sensitive to H2O2. Cells that were only deprived of glucose demonstrated no potentiation of injury, while cells subjected to mitochondrial inhibition with oligomycin alone also showed significant potentiation of oxidant injury. H2O2 consumption was not affected by metabolic inhibition. Conditions associated with mitochondrial inhibition consistently resulted in a decrease in intracellular pH. These experiments suggest that a synergism exists between metabolic inhibition and subsequent oxidant exposure.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Adenosine Triphosphate; Animals; Cattle; Cell Survival; Endothelium, Vascular; Glucose; Hydrogen Peroxide; Hydrogen-Ion Concentration; In Vitro Techniques; Ischemia; Models, Biological; Oligomycins; Potassium Cyanide; Reperfusion Injury