Radiation Oncology Department, Radiation and Cancer Biology Division, Washington University School of Medicine, St Louis, Missouri 63108, USA.
Int J Hyperthermia. 2010 Feb;26(1):67-78. doi: 10.3109/02656730903417947.
The goal of this study was to determine whether short-duration (15 s-3 min) high-temperature (50 degrees C) heat shocks inhibit the repair of DNA damage.
Cultured HeLa cells were used. DNA damage was measured after UV exposure or X-irradiation. Three methods were used to measure DNA damage: alkaline comet assay with the endonuclease, UVDE, for single strand breaks and UV photoproducts, antibodies specific for cyclo-pyrimidine dimers (CPD) or for 6-4 photoproducts (64PP), and the appearance-resolution of gamma-H2AX foci for DNA double strand breaks.
Heat shocks of 15-30 s at 50 degrees C inhibited repair of DNA damage after UV exposure or X-irradiation detected by the alkaline comet assay (after UV) or by persistence of gamma-H2AX foci (after X-rays). The phosphorylation of histone, H2AX, induced by 1 or 4 Gy of X-rays was inhibited in a time-dependent manner after 15-45 s at 52 degrees C. When the excision of UV-induced PP was measured, heat shocks of more than 60 s at 50 degrees C were required to show measurable inhibition.
Severe (50 degrees C) short-duration (15 s or greater) heat shocks inhibit repair of UV-induced DNA damage. The ability to detect the inhibitory effects of very short, 15-60 s, heat shocks was assay dependent. The comet assay could detect repair inhibition after a 15-s heat shock. Detection of DNA damage by specific antibodies could only detect repair inhibition after 1-3-min heat shocks. Using the gamma-H2AX foci method 30 s at 50 degrees C induced a significant delay in the repair of DNA damage after 1 Gy of X-rays.
本研究旨在确定短时间（15 秒至 3 分钟）高温（50°C）热休克是否会抑制 DNA 损伤的修复。
使用培养的 HeLa 细胞。紫外线照射或 X 射线照射后测量 DNA 损伤。使用三种方法测量 DNA 损伤：碱性彗星试验结合内切酶 UVDE 测量单链断裂和紫外线光产物，针对环嘧啶二聚体（CPD）或 6-4 光产物（64PP）的抗体，以及 γ-H2AX 焦点的出现分辨率测量 DNA 双链断裂。
50°C 下 15-30 秒的热休克抑制了紫外线照射或 X 射线照射后碱性彗星试验（紫外线后）或 γ-H2AX 焦点持续存在（X 射线后）检测到的 DNA 损伤修复。X 射线诱导的组蛋白 H2AX 磷酸化在 52°C 下 15-45 秒后呈时间依赖性抑制。当测量紫外线诱导的 PP 切除时，需要 50°C 下超过 60 秒的热休克才能显示可测量的抑制作用。
严重（50°C）短时间（15 秒或更长时间）的热休克抑制紫外线诱导的 DNA 损伤修复。检测非常短的 15-60 秒热休克抑制作用的能力取决于检测方法。彗星试验可以检测到 15 秒热休克后的修复抑制。特定抗体检测 DNA 损伤只能检测到 1-3 分钟热休克后的修复抑制。使用 γ-H2AX 焦点方法，50°C 下 30 秒会导致 1Gy X 射线后 DNA 损伤修复明显延迟。
