Quantitative analysis of γ-H2AX foci formation and dynamic changes in DNA double-strand breaks induced by X-ray radiation
10.3760/cma.j.issn.1004-4221.2018.03.015
- VernacularTitle:X射线诱导γH2AX焦点形成的定量分析及其致DNA双链断裂的动态变化
- Author:
Jun DONG
1
;
Chengtao WANG
;
Chun ZHANG
;
Yufeng REN
;
Bin OOYANG
;
Tian ZHANG
;
Zhenyu WANG
;
Li C. GLORIA
;
He FUQIU
;
Bixiu WEN
Author Information
1. 中山大学附属第一医院放射治疗科
- Keywords:
Non-homologous terminal connection;
DNA dependent protein kinase catalytic subunit;
γH2AX foci formation;
SUNE-1 cell line
- From:
Chinese Journal of Radiation Oncology
2018;27(3):303-308
- CountryChina
- Language:Chinese
-
Abstract:
Objective To quantitatively compare the γ-H2AX foci formation between DNA-PKcs+/+and DNA-PKcs-/-mouse embryonic fibroblast(MEF)cells,and to investigate the dynamic changes in DNA double-strand breaks(DSBs)in human nasopharyngeal carcinoma SUNE-1 cells exposed to X-ray radiation. Methods The expression of DNA-PKcs was determined by Western blot. The γ-H2AX foci formation induced by 5 Gy X-ray radiation was detected by cell immunofluorescence. The ImageJ software was used to quantitatively analyze the γ-H2AX foci formation. Results The expression of DNA-PKcs was silenced in DNA-PKcs-/-MEF cells and normal in DNA-PKcs+/+MEF cells. According to the dynamic analyses of the numbers of γ-H2AX foci/cell and γ-H2AX foci/mm2, a similar tendency was observed in DSB formation in DNA-PKcs+/+MEF cells, DNA-PKcs-/-MEF cells,and SUNE-1 cells exposed to X-ray radiation. A large number of γ-H2AX foci formed at 0.5-1.0 h after radiation. DSBs were repaired at 6 h after radiation in DNA-PKcs+/+MEF cells and 24 h after radiation in DNA-PKcs-/-MEF cells and SUNE-1 cells. The peak values of γ-H2AX foci/cell and γ-H2AX foci/mm2were observed at 1.0 and 0.5 h after radiation, respectively. Compared with DNA-PKcs+/+MEF cells, DNA-PKcs-/-MEF cells had different numbers of γ-H2AX foci/cell at 0.5, 1.0, 3.0, 6.0, and 12.0 h after radiation, as well as different numbers of γ-H2AX foci/mm2at 3.0, 6.0, and 12.0 h after radiation. Conclusions Quantitative measurement of the number of γ-H2AX foci/cell or γ-H2AX foci/mm2by cell immunofluorescence provides new insights into the quantitative and dynamic study of DSB damage and repair.
