南方医科大学学报2016，Vol.36Issue(4)：472-476,5.
S期H1299细胞热损伤后的延迟性DNA双链断裂
Delayed DNA double-strand break in S-phase H1299 cells after thermal damage
摘要
Abstract
Objective To study the pattern of DNA double-strand break (DSB) formation in S-phase cells after thermal damage and explore the mechanisms behind heat sensitivity of S-phase cells and delayed DSBs. Methods Flow cytometry was used to analyze the cell cycle arrest in H1299 cells exposed to thermal damage, and EdU incorporation assay was employed to evaluate the DNA replication capacity of the cells. The cells synchronized in S phase were obtained by serum starvation and DSBs were observed dynamically using neutral comet assay. Trypan blue dye exclusion technique was used to analyze the cell viability after thermal damage. Western blotting (WB) was used to detect the phosphorylation of ATM and DNA binding RAD18. Results The percentage of S-phase cells increased significantly after exposure of the cells to 45 ℃ for 1 h (P<0.01). The time-dependent variation pattern of EdU incorporation was similar to that of S-phase cell fraction. The comet tail began to appear only after incubation of the cells at 37℃for some time and the Olive tail moment (OTM) increased with prolonged incubation. Cell death remained low until 7.5 h after heat exposure of the S-phase cells and then increased rapidly. The phosphorylation of ATM first increased but then decreased drastically. In cells with heat exposure, DNA binding RAD18 was attenuated obviously compared that in non-exposed cells. Conclusion Thermal damage causes cell cycle arrest in S phase, and delayed fatal DSBs occur in the arrested cells due to persistent replication and DNA damage repair suppression, which are the possible cause of heat sensitivity of S-phase cells.关键词
H1299细胞/热损伤/细胞周期阻滞/DNA双链断裂Key words
H1299 cells/thermal damage/cell cycle arrest/DNA double-strand break引用本文复制引用
孙婷,丁为民,李凌,张彦..S期H1299细胞热损伤后的延迟性DNA双链断裂[J].南方医科大学学报,2016,36(4):472-476,5.基金项目
国家自然科学基金（81272407）；卫生部重大科技专项课题资助项目（W2009BX015） Supported by National Natural Science Foundation of China (81272407) （81272407）
