Mitochondrial dysfunction during hypoxia/reoxygenation and its correction by anaerobic metabolism of citric acid cycle intermediates

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Abstract
Kidney proximal tubule cells developed severe energy deficits during hypoxia/reoxygenation not attributable to cellular disruption, lack of purine precursors, the mitochondrial permeability transition, or loss of cytochrome c . Reoxygenated cells showed decreased respiration with complex I substrates, but minimal or no impairment with electron donors at complexes II and IV. This was accompanied by diminished mitochondrial membrane potential (ΔΨ m ). The energy deficit, respiratory inhibition, and loss of ΔΨ m were strongly ameliorated by provision of α-ketoglutarate plus aspartate (αKG/ASP) supplements during either hypoxia or only during reoxygenation. Measurements of 13 C-labeled metabolites in [3- 13 C]aspartate-treated cells indicated the operation of anaerobic pathways of αKG/ASP metabolism to generate ATP, yielding succinate as end product. Anaerobic metabolism of αKG/ASP also mitigated the loss of ΔΨ m that occurred during hypoxia before reoxygenation. Rotenone, but not antimycin or oligomycin, prevented this effect, indicating that electron transport in complex I, rather than F 1 F 0 -ATPase activity, had been responsible for maintenance of ΔΨ m by the substrates. Thus, tubule cells subjected to hypoxia/reoxygenation can have persistent energy deficits associated with complex I dysfunction for substantial periods of time before onset of the mitochondrial permeability transition and/or loss of cytochrome c . The lesion can be prevented or reversed by citric acid cycle metabolites that anaerobically generate ATP by intramitochondrial substrate-level phosphorylation and maintain ΔΨ m via electron transport in complex I. Utilization of these anaerobic pathways of mitochondrial energy metabolism known to be present in other mammalian tissues may provide strategies to limit mitochondrial dysfunction and allow cellular repair before the onset of irreversible injury by ischemia or hypoxia.