THE MICRONUCLEUS ASSAY - AN EVALUATION OF ITS USE IN DETERMINING RADIOSENSITIVITY IN-VITRO

The cytokinesis-block micronucleus assay was used to measure radiosensitivity in vitro in a panel of seven cell lines. Six of these cell lines were used to study the major parameters of this assay. We observed varying sensitivities following cytochalasin-B exposure. Treatment with 1 mu g/ml cytochalasin-B for 24 h reduced cell survival in four of the six cell lines by >60%, Cytochalasin-B concentration and post-irradiation culture time were both found to influence cell-response. In three cell lines (V39, V134 and HX142), a decrease in cytochalasin-B concentration (2-0.5 mu g/ml) resulted in an increase in the frequency of radiation-induced micronuclei per binucleate cell. In other cell lines, either the opposite (V7M, CHO-K1) or no effect (WiDr) was seen. A linear dose-response was observed between induced damage expressed as the frequency of micronuclei and radiation dose in all but one melanoma (V39) cell line. Evidence for radiation-induced division-delay, with the maximum frequency of binucleation in irradiated cultures occurring 24-48 h after that of controls, was only seen in two cell lines. Of particular note, and in contrast to some other published reports, was the lack of a general correlation between cell-response measured in the clonogenic and the cytokinesis-block micronucleus assays. Consideration of lethal lesions, determined from the clonogenic dose-response curve, with respect to micronucleus frequency showed a complex relationship, with one micronucleus per binucleate cell corresponding to a wide range of lethal lesions depending on the cell line. It has been postulated that the binucleate cell with no micronuclei may represent the surviving cell; however, we found no correlation between the slope of the frequency of these cells with respect to radiation dose and the clonogenic alpha slope. These observations should be considered prior to attempting to use the cytokinesis-block micronucleus assay to measure in vitro radiosensitivity in human tumour cells.