Reoxygenation of the hypoxic myocardium causes a mitochondrial complex I defect

Abstract

It is well established that reoxygenation of myocardial tissue after a period of hypoxia or ischaemia results in an increase in intracellular calcium [ 11 and the release of cytosolic contents as a consequence of cell lysis 121. As yet the mechanism and sequence of events involved in the so-called 'oxygen paradox' are unresolved. An insight into the mechanism of reoxygenation damage may be gained by investigating those cells which have not sustained lethal damage.Under normal circumstances the energy demands of active cardiac tissue are almost entirely satisfied by mitochondrial ATP synthesis. Hence the effect of myocardial reoxygenation o n mitochondrial function is of the greatest significance. Consequently, the effects of hypoxia and/or reoxygenation o n mitochondrial function has been investigated.Isolated rat hearts were subjected to retrograde perfusion by the Langendorff method with Krebs bicarbonate buffer. After an initial 5 min equilibration period of perfusion with 1 1 m~-glucose and gassing with 02/COz ( 19 : 1 ), the control, normoxic hearts were then gassed for a further 45 min with glucose-free buffer. Hypoxic hearts were those gassed with N2/C0, (10: 1) for 40 min with glucose-free buffer, while reoxygenated hearts were subject to a further 5 min of perfusion with O,/CO, (1Y:l). At the end of the perfusion, mitochondria were prepared by trypsin digestion. homogenization and differential centrifugation.The activity of the electron transport chain was determined by using a Clark-type electrode, the mitochondrial respiration rates being measured in the presence of both NAD-linked (pyruvate + malate) and FAD-linked (succinate) substrates. The activities of the individual respiratory chain complexes isolated from the membrane by freeze/thawing were determined by assaying NADH-cytochrome c reductase activity (complex 1-111) and succinate-cytochrome c reductase activity (complex 11-111). Table 1 shows that there is a 44% decrease in the state 3 respiration rate with NAD-linked substrates, but no reduction in the succinate-linked rate, in mitochondria isolated from hearts subjected to hypoxia and reoxygenation, but not hypoxia alone. The respiratory control ratios are reduced after reoxygenation, but without an increase in state 4 respiration (data not shown), which suggests that membrane integrity is unaffected. The reduction in complex I activity is confirmed by the enzyme assay data in Table 1. Reoxygenated isolated hypoxic heart mitochondria do not show the complex I defect, suggesting that the mechanism involves the intervention of the whole myocyte or tissue. Isolated adult cardiomyocytes subject to hypoxia and reoxygenation have been shown not to undergo cell lysis 131, but preliminary studies have shown that they do exhibit the complex I defect. This suggests that the whole tissue is not necessary for the defect t o be expressed, and that the relationship of the mitochondrial defect and cell lysis may be indirect.Evidence has been presented that the mitochondrial calcium uptake inhibitor Ruthenium...

DJ‐1 plays an obligatory role in the cardioprotection of delayed hypoxic preconditioning against hypoxia/reoxygenation‐induced oxidative stress through maintaining mitochondrial complex I activity

“…Mitochondria is one of the major organelles that regulates the ROS production, whereas mitochondrial complex I is a main site of ROS production in mitochondria . Substantial evidence is accumulating to suggest that the defect of mitochondrial complex I is a critical determinant of increased mitochondrial ROS generation and subsequent oxidative stress following myocardial I/R injury . Therefore, preventive and therapeutic methods targeting mitochondrial complex I may be more efficient against H/R‐induced oxidative stress.…”

Endothelial cell respiration is affected by the oxygen tension during shear exposure: role of mitochondrial peroxynitrite

“…ONOO Ϫ -mediated inactivation of complex I increases O 2 ⅐ Ϫ production from that location, and O 2 ⅐ Ϫ generated by complex I is released exclusively into the matrix (16,51,(63)(64)(65). Animal I/RP models showed that, upon RP, the ETC complex activities are reduced, complex I is the main source of mitochondrial ROS (2,31,54), and NO derived from endothelial NOS (eNOS) is the one responsible for ONOO Ϫ formation, inhibition of complex I activity, and suppression of tissue O 2 consumption (84).…”

Mechanism of Ischemia and Reperfusion Injury to the Heart: From the Viewpoint of Nitric Oxide and Mitochondria

“…The hallmarks of cardiac IR injury are found to occur on reperfusion of the myocardium. In terms of the mitochondria, reperfusion injury affects the oxidative phosphorylation (Ox-Phos) pathway, including the ETC, [53][54][55] adenine nucleotide translocase (ANT), 56,57 and Krebs cycle 58,59 enzymes. In addition to Ox-Phos, reperfusion in-jury also leads to cardiolipin oxidation, 55,60,61 the induction of a large proton leak across the mitochondrial inner membrane, 41,62 Ca 2+ overload, overproduction of ROS, PTP opening, and cell death.…”

“…In addition to Ox-Phos, reperfusion in-jury also leads to cardiolipin oxidation, 55,60,61 the induction of a large proton leak across the mitochondrial inner membrane, 41,62 Ca 2+ overload, overproduction of ROS, PTP opening, and cell death. [62][63][64][65][66][67][68][69][70][71][72][73] A majority of these observations have been made in mitochondria isolated from hearts after reperfusion, meaning that the time course of mitochondrial damage during reperfusion injury is difficult to study (because mitochondrial isolation typically takes 1∼2 h).…”