Short-term culture and γH2AX flow cytometry determine differences in individual radiosensitivity in human peripheral T lymphocytes
Short-term culture and γH2AX flow cytometry determine differences in individual radiosensitivity in human peripheral T lymphocytes
复制标题
DOI：
10.1002/em.20273
复制
发表时间：
2007-01-01
影响因子：
2.8
通讯作者：
Kusunoki, Yoichiro
中科院分区：
文献类型：
--
作者：
Hamasaki, Kanya;Imai, Kazue;Kusunoki, Yoichiro
组蛋白H2AX是组蛋白H2A的一个亚家族，其被磷酸化并在DNA双链断裂位点处形成蛋白质修复灶，以响应遗传毒性损伤，如电离辐射。这个过程被认为在DNA损伤的修复中起着关键作用。在这项研究中，我们建立了一个流式细胞术（FCM）系统，用于测量辐射诱导的磷酸化组蛋白H2AX（γ H2AX）在培养的人T淋巴细胞，以评估个人辐射敏感性在体外。照射短期（类似于7天）培养的T淋巴细胞表现出显着的个体间，但不interexperimental，差异γ H2AX的细胞含量6小时后，4戈伊的X射线照射在三个独立的实验，使用外周血淋巴细胞从6个健康供体。然而，这些差异在未培养的淋巴细胞或长时间培养的淋巴细胞（类似于13天）中并不明显。个体淋巴细胞中γ H2AX病灶形成的变化是可重复的，培养7天后差异达到约1.5倍。因此，基于FCM的γ H2AX测量似乎反映了辐射淋巴细胞内的DNA损伤的时间过程和量。此外，我们证实，残留淋巴细胞亚群的差异不涉及个体的放射敏感性。这些结果表明，FCM为基础的γ H2AX检测培养的T淋巴细胞可能是有用的快速和可靠的评估参与DNA损伤修复的个人辐射敏感性。
Histone H2AX, a subfamily of histone H2A, is phosphorylated and forms proteinaceous repair foci at the sites of DNA double-strand breaks in response to genotoxic insults, such as ionizing radiation. This process is believed to play a key role in the repair of DNA damage. In this study, we established a flow cytometry (FCM) system for measuring radiation-induced phosphorylated histone H2AX (gamma H2AX) in cultured human T lymphocytes to evaluate individual radiation sensitivity in vitro. Irradiation of short-term (similar to 7 days) cultured T lymphocytes exhibited significant interindividual, but not interexperimental, differences in the cellular content of gamma H2AX 6 hr after 4 Gy of X-irradiation in three independent experiments using peripheral blood lymphocytes from six healthy donors. However, these differences were not as marked in uncultured lymphocytes, or lymphocytes that were cultured for a prolonged period (similar to 13 days). The variation of gamma H2AX focus formation in lymphocytes of individuals was reproducible, with differences reaching about 1.5-fold following 7 days of culture. Therefore, the FCM-based gamma H2AX measurement appeared to reflect both the temporal course and the amount of DNA damage within the irradiated lymphocytes. Further, we confirmed that the differences in residual lymphocyte subsets were not involved in individual radiosensitivity. These results suggest that the FCM-based gamma H2AX assay using cultured T lymphocytes might be useful for the rapid and reliable assessment of individual radiation sensitivity involved in DNA damage repair.