Photosensitization and split-dose recovery in cultured human urinary bladder carcinoma cells containing...

The photosensitization and survival recovery of cultured EJ human urinary bladder carcinoma cells containing nonexchangeable hematoporphyrin derivative (HPD) were studied. Cultures were incubated at 37 degrees C in growth medium supplemented with HPD (50 micrograms/ml) and 5% fetal bovine serum for 12 h followed by incubation in HPD-free medium containing 5% fetal bovine serum for 9 or 18 h. The levels of porphyrin remaining in the cells (termed the "nonexchangeable" intracellular porphyrin component) were not significantly different at these times, and as a result sensitivities to broad-band red light (greater than 580 nm) were also identical. Shouldered survival curves were obtained in each case, indicating the ability to accumulate sublethal photodamage. Recovery from photosensitized damage using a split-dose technique was examined. Single, attached, asynchronously growing cells containing nonexchangeable HPD (12 h HPD uptake plus 9 h in porphyrin-free medium) were exposed to red light (1.2 kJ/sq m) and, after various intervals at 37 degrees C in the dark, a second dose of 1.2 kJ/sq m. Survival rapidly increased and reached a maximum at about 9 h between light doses. Analysis of dose-response curves revealed a partial reappearance of the curve shoulder (Dq = 0.22 kJ/sq m) and a markedly reduced curve slope (D0 = 0.82 kJ/sq m) for fractionated irradiations with a 9-h interval in comparison with graded, single light exposures (Dq = 0.48 kJ/sq m; D0 = 0.41 kJ/sq m). These observations suggest that the cells developed an increased tolerance to photosensitized damage after prior HPD-light treatment. No significant change in intracellular HPD levels between irradiations was detected, indicating that the increased survival was not due to a loss of sensitizer from inside the cells. These results demonstrate that EJ cells accumulate and recover from HPD-sensitized photodamage; analogous to the accumulation and recovery from sublethal damage (Elkind recovery) in other mammalian cultures treated with ionizing radiation.