Takahashi Akihisa, Yamakawa Nobuhiro, Kirita Tadaaki, Omori Katsunori, Ishioka Noriaki, Furusawa Yoshiya, Mori Eiichiro, Ohnishi Ken, Ohnishi Takeo
Department of Biology, School of Medicine, Nara Medical University, Nara, Japan.
J Radiat Res. 2008 Nov;49(6):645-52. doi: 10.1269/jrr.08007. Epub 2008 Nov 6.
To identify the repair dynamics involved in high linear energy transfer (LET) radiation-induced DNA damage, phospho-H2AX (gammaH2AX) foci formation was analyzed after cellular exposure to iron ions (Fe-ions, 500 MeV u(-1), 200 KeV microm(-1)). The foci located at DNA damage sites were visualized using immunocytochemical methods. Since H2AX is phosphorylated at sites of radiation-induced double strand breaks (DSB), gammaH2AX foci were used to detect or illuminate tracks formed by DSB after exposure to various doses of ionizing radiation. Additional DSB-recognition proteins such as ATM phospho-serine 1981, DNA-PKcs phospho-threonine 2609, NBS1 phospho-serine 343 and CHK2 phospho-threonine 68 all co-localized with gammaH2AX at high LET radiation induced DSB. In addition, Fe-ion induced foci remained for longer times than X-radiation induced foci. These findings suggest that Fe-ion induced damage is repaired more slowly than X-radiation induced damage, possibly because Fe-ion induced damage or lesions are more complex or extensive. Antibodies for all these phosphorylated DNA DSB recognition proteins appear to be very effective for the detection and localization of DSB.
为了确定高传能线密度（LET）辐射诱导的DNA损伤所涉及的修复动力学，在细胞暴露于铁离子（Fe离子，500 MeV u(-1)，200 KeV μm(-1)）后，分析了磷酸化H2AX（γH2AX）焦点的形成。使用免疫细胞化学方法观察位于DNA损伤位点的焦点。由于H2AX在辐射诱导的双链断裂（DSB）位点被磷酸化，因此γH2AX焦点被用于检测或显示暴露于不同剂量电离辐射后由DSB形成的轨迹。其他DSB识别蛋白，如ATM磷酸化丝氨酸1981、DNA-PKcs磷酸化苏氨酸2609、NBS1磷酸化丝氨酸343和CHK2磷酸化苏氨酸68，在高LET辐射诱导的DSB处均与γH2AX共定位。此外，Fe离子诱导的焦点比X射线诱导的焦点持续时间更长。这些发现表明，Fe离子诱导的损伤比X射线诱导的损伤修复得更慢，这可能是因为Fe离子诱导的损伤或病变更复杂或更广泛。所有这些磷酸化DNA DSB识别蛋白的抗体似乎对DSB的检测和定位非常有效。
