cytochrome-c-t and 2-2-dimethyl-5-hydroxy-1-pyrrolidinyloxy

Glutamate-cysteine ligase (GCL), a rate-limiting enzyme for glutathione (GSH) synthesis, is composed of catalytic and modifier subunits. This study examined the pathogenic role of GCL modifier subunits (GCLM) in myocardial ischaemia-reperfusion (I/R) injury using mice lacking the GCLM (GCLM(-/-)).. The GCLM(-/-)mice had an increase in myocardial I/R injury and apoptosis in ischaemic myocardium compared with GCLM(+/+) mice. There was a decrease in mitochondrial glutathione (GSH) levels in ischaemic myocardium that was more pronounced in GCLM(-/-) mice than in GCLM(+/+) mice (12 vs. 55% of baseline GCLM(+/+), respectively). The ESR signal intensity of the dimethyl-1-pyrroline-N-oxide-hydroxyl radical adducts in ischaemic myocardium was higher in GCLM(-/-) mice than in GCLM(+/+) mice. Hypoxia-reoxygenation induced greater mitochondrial damage in cultured cardiomyocytes from GCLM(-/-) mice than from GCLM(+/+) mice, as evidenced by a reduced membrane potential and increased protein carbonyl content in isolated mitochondria, together with enhanced cytochrome c translocation into the cytosol. Administration of GSH ethyl-ester attenuated myocardial I/R injury and reversed the mitochondrial damage in parallel with the mitochondrial GSH restoration in the myocardium or the cardiomyocytes of GCLM(-/-) mice.. GCLM(-/-) mice were susceptible to myocardial I/R injury partly through an increased vulnerability of mitochondria to oxidative damage owing to mitochondrial GSH reduction.

Topics: Animals; Caspase 3; Caspase 9; Cells, Cultured; Cyclic N-Oxides; Cytochromes c; Echocardiography; Glutamate-Cysteine Ligase; Glutathione; Male; Mice; Mice, Inbred C57BL; Mice, Mutant Strains; Mitochondria; Myocardial Infarction; Myocardial Reperfusion Injury; Myocardium; Myocytes, Cardiac; Oxidative Stress; Reactive Oxygen Species; RNA, Messenger; Ventricular Function, Left

An electron paramagnetic resonance (EPR) signal characteristic of the 5,5'-dimethyl-1-pyrroline-N-oxide (DMPO)-OH spin adduct, which is formed from the reaction of DMPO with superoxide radicals generated by xanthine oxidase-mediated reaction, was significantly reduced by the cadaverine or Escherichia coli Mn-containing superoxide dismutase (MnSOD). Likewise, cytochrome c reduction by superoxide was inhibited by cadaverine, and the inhibition level increased in proportion to the level of cadaverine. The cadA mutant of Vibrio vulnificus, which does not produce cadaverine because of the lack of lysine decarboxylase, exhibits less tolerance to superoxide stress in comparison with wild type. The results indicate that cadaverine scavenges superoxide radicals, and protects cells from oxidative stress.

Topics: Antiporters; Bacterial Proteins; Cadaverine; Cyclic N-Oxides; Cytochromes c; Electron Spin Resonance Spectroscopy; Genes, Bacterial; Mutation; Oxidation-Reduction; Oxidative Stress; Spin Labels; Superoxides; Vibrio vulnificus