Some characteristics of hyperoxia-adapted HeLa cells. A tissue culture model for cellular oxygen tolerance

By culturing HeLa cells at stepwise increased oxygen tensions over a prolonged period of time (approximately 21 months) we selected a substrain capable of growing under 80% O2/19% N2/1% CO2, an oxygen level that is lethal to normal HeLa cells, adapted to 20% O2/79% N2/1% CO2. The 80% O2-adapted cells exhibited the following characteristics. At the ultrastructural level an abnormal mitochondrial morphology was observed: compared to normal cells, mitochondria of the hyperoxia-adapted cells exhibited a 3-fold larger mean profile area in sections and were slightly decreased in number; the relative mitochondrial volume was increased 2-fold, whereas the size of both cell types was the same. Mitochondrial matrix appeared less dense in the hyperoxia-adapted cells; no structural damage was detected. Compared to the 20% O2-adapted cells O2 consumption per cell was approximately 40% decreased in the 80% O2-adapted cells. Under hyperoxic conditions 20% O2-adapted and 80% O2-adapted cells exhibited very similar cyanide-resistant respiration rates (0.16 +/- 0.04 and 0.15 +/- 0.02 fmoles/cell/minute, respectively), suggesting that the increased O2 tolerance of the 80% O2-adapted cells was not due to a decreased cellular production of activated oxygen species at hyperoxia. Cellular levels of the enzymes directly involved in protection against activated oxygen species, i.e., superoxide dismutases, catalase, and glutathione peroxidase, were normal or slightly below normal in the 80% O2-adapted cells, implying that these enzymes were of no significance for the increased O2 tolerance. In addition, the specific activity of glucose-6-phosphate dehydrogenase, a key enzyme for cellular production of NADPH, was not related to the degree of O2 tolerance. Our results suggest that the increased O2 tolerance of the 80% O2-adapted cells is neither based on cellular properties controlling the formation or removal of intracellular activated oxygen species nor on the cellular capacity to repair or replace damaged cellular components. We speculate that the increased O2 tolerance is largely due to a genetically determined increased resistance of oxygen-sensitive cellular targets.