cytochrome-c-t and Pulmonary-Edema

Reduced glutathione (GSH) has been shown to improve pulmonary graft preservation. Mitochondrial dysfunction is regarded to be the motor of ischemia-reperfusion injury (IR) in solid organs. We have shown previously that IR induces pulmonary mitochondrial damage. This study elucidates the impact of GSH preconditioning on the integrity and function of pulmonary mitochondria in the setting of warm pulmonary IR.. Wistar rats were subjected to control, sham, and to two-study-group conditions (IR30/60 and GSH-IR30/60) receiving IR with or without GSH preconditioning. Rats were anesthetized and received mechanical ventilation. Pulmonary in situ clamping followed by reperfusion generated IR. Mitochondria were isolated from pulmonary tissue. Respiratory chain complexes activities (I-IV) were analyzed by polarography. Mitochondrial viability (Ca2+-induced swelling) and membrane integrity (citrate synthase assay) were determined. Subcellular-fractional cytochrome C-content (Cyt C) was quantified by enzyme-linked immunosorbent assay (ELISA). Mitochondrial membrane potential (ΔΨm) was analyzed by fluorescence-activated cell sorting (FACS) after energizing and uncoupling. Inflammatory activation was determined by myeloperoxidase activity (MPO), matrix-metalloproteinase 9 (MMP-9) activity by gel zymography.. Pulmonary IR significantly reduced mitochondrial viability in combination with ΔΨm hyper-polarization. GSH preconditioning improved mitochondrial viability and normalized ΔΨm. Cyt C was reduced after IR; GSH protected from Cyt C liberation. Respiratory chain complex activities (I, II, III) declined during IR; GSH protected complex II function. GSH also protected from MMP-9 and neutrophil sequestration (P>.05).. GSH preconditioning is effective to prevent mitochondrial death and improves complex II function during IR, but not mitochondrial membrane stability. GSH-mediated amelioration of ΔΨm hyper-polarization appears to be the key factor of mitochondrial protection.

Topics: Animals; Apoptosis; Calcium; Cytochromes c; Disease Models, Animal; Drug Evaluation, Preclinical; Electron Transport; Glutathione; Granulocytes; Ischemic Preconditioning; Lung; Matrix Metalloproteinase 9; Membrane Potential, Mitochondrial; Mitochondria; Mitochondrial Diseases; Oxygen Consumption; Pulmonary Edema; Rats; Rats, Wistar; Reperfusion Injury

This study examines the impact of glycine (Gly) preconditioning on ischemia reperfusion (IR)-induced pulmonary mitochondrial injury to research the previously, in pig lungs, demonstrated Gly-dependent amelioration of pulmonary IR injury. IR injury was induced in rat lungs by 30 min pulmonary hilum clamping followed by 60 min reperfusion time. Rats were subjected to controls, shams and two study groups (IR30/60, Gly-IR30/60) receiving 37.5 mg Gly i.v. or not before IR induction. The wet/dry-weight ratio, mitochondria viability (MV), membrane integrity (MI), respiratory chain complex (RCC) activities, mitochondrial membrane potential (ΔΨm) and cytochrome C (Cyt C) content were analysed. In IR30/60, RCC and MV were impaired; Cyt C loss and MI combined with matrix metalloproteinase-9 (MMP-9) activation and ΔΨm alteration were observed when compared with controls. In Gly-IR30/60, complex II function and mitochondrial viability were protected during IR, and MMP-9 activation combined with tissue-water content accumulation and ΔΨm alteration were ameliorated. Cyt C loss, mitochondrial membranes damage, tissue GSH oxidation or neutrophil sequestration was not extenuated in Gly-IR30/60. Gly ameliorates IR-associated mitochondrial dysfunction and decay of viability and normalizes ΔΨm but does not protect from Cyt C liberation and mitochondrial membrane damage. Our data suggest that the previously described effect of Gly preconditioning results at least partially from mitochondrial protection. A dose-finding study is necessary to improve results of Gly preconditioning.

Topics: Animals; Apoptosis; Cytochromes c; Cytoprotection; Disease Models, Animal; Electron Transport Chain Complex Proteins; Enzyme Activation; Glutathione; Glycine; Lung; Male; Matrix Metalloproteinase 9; Membrane Potential, Mitochondrial; Mitochondria; Neutrophil Infiltration; Oxidation-Reduction; Protective Agents; Pulmonary Edema; Rats; Rats, Wistar; Reperfusion Injury; Time Factors