Differences in heating methods may account for variation in reported effects on γH2AX focus formation

Previously we have shown that heat shock cannot induce DNA damage in several mammalian cell lines, using γH2AX foci formation as an indicator for DNA damage. However, there have also been reports showing heat shock can induce DNA damage. In an effort to clarify why such differences exist among different studies, we examined the effects of five different heating methods, e.g., (1) heat block; (2) submerged in water bath; (3) half-submerged in water bath; (4) pre-heated medium; and (5) heating oven, on γH2AX foci formation in human amnion FL cells and Chinese hamster CHL cells. It was found that during a 24 h period, heating at 42 °C for 30 min showed no significant cytotoxicity, while 45 °C exhibited various degrees of cytotoxicity. The ability of these five methods to induce γH2AX foci was quite distinct, with only heat block inducing γH2AX foci at both 42 °C and 45 °C in both FL and CHL cells. The two cell types also responded differently to the different heating methods, with CHL being more sensitive to heat shock. Taken together, it is clear that certain heating methods can induce γH2AX foci formation; on the other hand, their effects differ in thermo-tolerant and non-tolerant cells. And the difference in γH2AX foci formation observed in different reports most likely is a consequence of the different heating methods used in the study. However, expression of γH2AX may not be used as a surrogate of cell killing.