UBC Theses and Dissertations
Respiration induced oxygen gradients in cultured mammalian cells Fengler, John Josef Paul
Oxygen is known to sensitize X-irradiated cells to lethal radiation damage. At low ambient oxygen tensions, however, the molecular mechanisms of the sensitization process and the metabolic requirements of the cell may be forced to compete for the cellular oxygen supply. The effect of cell respiration on the availability of intracellular oxygen during irradiation was consequently investigated by comparing the radiosensitivities of respiring and non-respiring cells. Cultured mammalian cells were irradiated in single cell suspensions and thin film monolayers at respiration inhibiting (4°C) and at normal cell culturing (37°C) temperatures. Due to oxygen equilibration and radiolytic depletion problems, the results of the suspension culture experiments were inconclusive. By subsequently analyzing the diffusive mass transfer of oxygen in the suspension medium, the stirrer flask was determined to be an inappropriate culture vessel in which to irradiate cells at constant low oxygen concentrations. A thin film cell culture system in which the oxygen concentrations to which the cells were exposed during irradiation could be more accurately controlled was then developed. A comparison of the oxygen enhanced radiosensitivities of the respiring and non-respiring cells in thin film monolayers suggested that the metabolic depletion of oxygen at low oxygen tensions has a significant effect on the local and intracellular oxygen distribution. These effects are representative of those that would be produced if respiration induced oxygen gradients existed inside and immediately around respiring cells. The magnitude of the differential radiosensitivities was found to be dependent on cell shape and to have values that agreed very well with theoretical predictions based on the existence of such gradients.
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