Quantitative detection of γ-radiation-induced DNA double-strand breaks using γ-H2AX foci in mouse organs, lymphocytes and scid lymphoma cell line

一些生物标志物，如突变、染色体畸变、微核、集落形成等，已被用来测量辐射剂量。然而，需要大量的动物或细胞来测量由辐射引起的如此低频率的事件。为了更准确和灵敏地测量辐射损伤，最好检测辐射后细胞中的第一或最早事件(DNA双链断裂)，特别是在修复缺陷的细胞或动物中。小鼠(C.B17株)和SCID淋巴瘤细胞(来源于C.B17-SCID小鼠)用~(137)Cs~(γ)射线(0.1、0.2、0.5、1、2和4Gy射线)照射。通过测量细胞内γ-H_2AX斑点的信号强度或计数细胞内的荧光信号数，对SCID淋巴瘤细胞、C.B17外周淋巴细胞和器官中的DNA双链断裂进行定量。γ-H_2AX焦点在照射后3min内出现，60min达到最大荧光强度。在淋巴细胞、淋巴瘤细胞和小鼠器官(脑、肝、肾、睾丸)的细胞中，γ-H_2AX病灶的数量与辐射剂量之间的关系在所研究的整个剂量范围内是线性的。这些结果表明，用γ-H_2AX焦点定量DNA双链断裂是检测小鼠高剂量和低剂量辐射损伤的有用的生物剂量计。有趣的是，在来自小肠的绒毛细胞中，没有发现照射后γ-H_2AX焦点的增加。与γ-H_2AX磷酸化相关的基因本应在器官的所有细胞中表达，以修复辐射和化学物质等环境影响引起的自发DNA双链断裂，然而，似乎这些必要的系统并不是在所有器官中都起作用。

Several biological markers, mutation, chromosome aberration, microneuclei, colony formation, etc. have been used to measure radiation doses. However, enormous numbers of animals or cells are required to measure such low frequent events induced by radiations. To measure the radiation damage more precisely and sensitively, it is better to detect the first or earliest events in cells after irradiation (DNA double-strand breaks), especially in repair deficient cells or animals. Mice (C.B17 strain) and scid lymphoma cells (from C.B17-scid mice) were exposed to ^<137>Csγ-rays (0.1, 0.2, 0.5, 1, 2 and 4 Gy). Quantitation of DNA double-strand breaks in scid lymphoma cells, C.B17 peripheral lymphocytes, and organs was carried out by measuring signal intensity or counting fluorescence signal number of γ-H2AX foci in cells. The γ-H2AX foci appeared within 3 min after irradiation and reached a maximum fluorescence in 60 min. In lymphocytes, lymphoma cells, and cells from the mouse organs (brain, liver, kidney, testis), the relationships between the numbers of γ-H2AX foci and radiation dose was found to be linear over the entire dose range studied. These results suggest that quantitation of DNA double-strand breaks by the γ-H2AX foci is an useful biodosimeter for detecting high and low dose radiation damage in mice. Intriguingly, in the villi cells from the small intestine, no increase of γ-H2AX foci was found after irradiation. Genes related to the γ-H2AX phosphorylation are supposed to express in all cells of organs to repair spontaneous DNA double-strand breaks caused environmental effects, e.g. radiation and chemicals, however, it seems that theses essential systems are not working in all organs.