Expression and Subcellular localization of DNA-PK in CNE1 and CNE2 Nasopharyngeal Carcinoma Cell Lines with different radiosensivity

摘要

Objective: Radiosensitivity is mainly determined by the number of DNA double-strand breaks (DSBs) induced by irradiation and the extent of its repair. The DNA-PK complex formation is one of the major pathways by which to repair the mammalian cells DSBs induced by ionizing radiation. This study was designed to elucidate whether there have some relationship between the expression and localization of Ku70, Ku80 and DNA-PKcs protein and the radiosensitivity of Nasopharyngeal Carcinoma cell lines CNE1/CNE2 with different radiosensitivity. Material and methods: Immumnohistochemistry was performed to detect the native subcellular localization of Ku70/Ku80/DNA-PKcs in NPC cells lines CNE1 and CNE2. Western-blot was designed to determine the expression of Ku protein in total extract of CNE1 and CNE2 and a semi-quantitative assay of protein expression was performed to estimate the optic density (OD) value of each band using automatic image analysis system. Result: Immumnohistochemistry suggest that Ku70/Ku80/DNA-PKcs primarily located in the nucleus, DNA-PKcs mainly in the nucleolus of CNE1 and CNE2. Western blotting and semi-quantitative assay ofprotein expression show that the integral optical density(IOD) of Ku70 protein in two cells is 22.03 ± 7.56 and 19.98 ± 6.04 respectively, and t=0.021, P > 0.05; While the integral optical density of Ku80 protein in two cells is 33.44±12.87 and 28.98 ± 9.24 respectively, t=0.24, P > 0.05; and the integral optical density of DNA-PKcs protein in two cells is 45.03 ± 1.77 and 40.87±4.19, t=1.58, P > 0.05. It suggested that the basic expression of Ku70/Ku80/DNA-PKcs had no statistic discrepancy between the different radiosensitive NPC cell lines CNE1 and CNE2. Conclusions: It indicates that there maybe no obviously correlation between the subcellular location and basic expression of DNA-PK protein and the variation of radiosensitivity in NPC cell lines CNE1 and CNE2. So we presume the difference of radiosensitivity between CNE1 and CNE2 may be on account of some other factors than subcellular localization and basic expression of DNA-PK.