Hyperoxia-induced clonogenic killing of HeLa cells associated with respiratory failure and selective inactivation of Krebs cycle enzymes

Cellular intoxication by elevated concentrations of O2 may be considered as a model for accelerated cellular aging processes resulting from excessive free radical production by normal metabolic pathways. We describe here that exposure of HeLa cell cultures to 80% O2 for 2 days causes progressive growth inhibition and loss of reproductive capacity. This intoxication was correlated with inhibition of cellular O2 consumption and inactivation of 3 mitochondrial flavoproteins, i.e., partial inactivation of NADH and succinate dehydrogenases and total inactivation of alpha-ketoglutarate dehydrogenase. As alpha-ketoglutarate dehydrogenase controls the influx of glutamine/glutamate into the Krebs cycle, which is the major pathway for oxidative ATP generation in HeLa cells, the inactivation of alpha-ketoglutarate dehydrogenase was expectedly correlated with a net fall in glutamine/glutamate utilization. Furthermore, a simultaneous increase in glucose consumption and lactate production was observed, indicating that the cellular response to respiratory failure is to generate more ATP from glycolysis. In spite of this response, extensive depletion of ATP was observed. Thus, hyperoxia-induced growth inhibition and loss of clonogenicity seem to be due primarily to an impairment of mitochondrial energy metabolism resulting from inactivation of SH-group-containing flavoprotein enzymes localized at or near the inner mitochondrial membrane. These observations may be relevant for theories implicating loss of mitochondrial function as a prime factor in the aging process.