Abstract:
The aim of this thesis is to evaluate if heat causes DNA double strand breaks (DSBs). There were several studies reported that heat increases of γ-H2AX foci, the serine 139 phosphorylated form of histone H2AX,, one of the earliest repair responses to DSBs. Nevertheless, there were no additional consequence of DSBs, such as apoptosis, was reported. Furthermore, heat did not demonstrate positive comet assay. Therefore, no actual evidence of DSB was detectable. Therefore, heat may induce γ-H2AX foci by other mechanisms besides DSBs or heat may induce a low number of DSBs. Consequently, the comet assay was negative and the consequences of heat induced DSBs were distinct from radiation. This thesis applied IRS-EDSB-LMPCR to compare the number of endogenous DSBs (EDSBs) between normal and heat induced genomes. Interspersed repetitive sequences (IRS) are randomly distributed. Hence, IRS-EDSB-LMPCR is a sensitive technique to measure rare and randomly distributed DSBs. We tested several epithelial and hematologic cell lines including, HeLa (cervical cancer), HN8, HN17 (Head and neck), Daudi (B lymphoblast), Molt4 (T lymphoblast) and Hacat (normal cell line) and found significant increase in EDSBs by heat in HeLa, p <0.001, and WSU-HN8, p<0.05. Interestingly, HeLa and WSU-HN8 possess the most genome-wide hypomethylation when compared with the others. This may suggest that heat increased EDSBs in cell may be correlate with chromatin structure. This hypothesis was proved by tricostatin A (TSA) treatment. Remarkably increased in heat induced EDSBs was found when TSA treated cells were heated, P<0.05.
