Cell-cycle dependence of heat-induced interphase death in mouse L5178Y cells

Cell lysis and eosin staining were observed in L5178Y cells within the first 3 h of post-hyperthermia incubation at 37 degrees C, after which both leveled to a plateau. Lysis and eosin staining were proportional to the severity of heat in asynchronous cells, whereas it was maximum in the most heat-sensitive M phase, intermediate in S, and least in heat-resistant G1 for the same heat treatment. Further, leakage of labeled [3H]thymidine and a decrease in radioactivity retained within heated cells coincided with an increase in eosin staining, indicating that the dye uptake was due to membrane damage. It was presumed that the eosin-stained fraction represented dead cells. The percentage eosin-stained cells reached a plateau, and this level was used to determine survival; when the results were compared with those obtained by the colony formation method, they were identical. By comparing the two survival assay methods we concluded that cell death after hyperthermia in L5178Y cells is mainly by interphase death in all phases of the cell cycle. The reasons for this conclusion are that a reduction in survival could be detected within one generation of L5178Y cells by the eosin staining method, and the survival values obtained by this method were identical to those obtained by the colony formation method.