To verify whether enhanced substrate-level phosphorylation increases viability and adenosine 5′-triphosphate (ATP) content of cells with neuropathy, ataxia, and retinitis pigmentosa/maternally inherited Leigh syndrome (NARP/MILS) mitochondrial DNA mutations and ATP synthase dysfunction.
We used cell lines “poisoned” with oligomycin, the specific inhibitor of ATP synthase, and “natural” models, including transmitochondrial human cell lines (cybrids) harboring 2 different pathogenic mutations associated with the NARP/MILS phenotypes.
Main Outcome Measures
Cell survival, morphology, and ATP content.
When normal human fibroblasts cultured in glucose-free medium were forced to increase energy consumption by exposure to the ionophore gramicidin or were energy challenged by oligomycin inhibition, their survival at 72 hours was 5%, but this increased to 70% when the medium was supplemented with α-ketoglutarate/aspartate to boost mitochondrial substrate-level phosphorylation. Homoplasmic cybrids harboring the 8993T→G NARP mutation were also protected from death (75% vs 15% survival at 72 hours) by the supplemented medium and their ATP content was similar to controls.
These results show that ATP synthase–deficient cells can be rescued by increasing mitochondrial substrate-level phosphorylation and suggest potential dietary or pharmacological therapeutic approaches based on the supplementation of α-ketoglutarate/aspartate to patients with impaired ATP synthase activity.