The Expression Of MRE11 Protein In CNE1 Cells After X-ray Irradiation

DNA break is always considered to be a cause of tumor radiotherapy failures. Ionizing radiation causes DNA damage, including deoxyribose change, base changes, DNA strand breaks, DNA double-strand breaks, formation of DNA-DNA crosslinking and other DNA-protein crosslinking. In which double-stranded DNA damage (double-stranded breaks, DSBs) is the most serious injury, leading to cell proliferation have lost the ability to split the death is caused by ionizing radiation and the main mechanism of tumor cell death. In normal cells, DNA double strand damage repair can enhance the radiation resistance of normal tissues in the radiation target, but in cancer cells, may lead to local recurrence and metastasis. Repair mechanisms include homologous recombination repair and non-homologous end joining.Studies show that the MRE11-RAD50-NBS1 complex in the repair of DSBs play an important role.The MRE11 protein has four amino-terminal phosphoesterase motifs and two DNA-binding motifs. Biochemical experiments have shown that the phosphoesterase domain of MRE11 functions as both a single-and double-stranded(ds) DNA endo-nuclease, as well as a 3'-5'dsDNA exonuclease, to recognize and bind to DSBs Department, and RAD50, NBS1 interact together to complete repair. The study, MRE11 expression by X-ray irradiation in tumor cells, can reveal new avenues of tumor radiotherapy, and research the passway regulated the expression of MRE11 can also provide a new target for tumor therapy.Objective:The study designed to determine the expression of MRE11 protein in nasopharyngeal carcinoma(NPC) cell lines CNE1. The expression changes of MRE11 mRNA after irradiation by OGy,2Gy,4Gy,6Gy,8Gy different doses in CNE1 cell lines. CNE1 cell lines were irradiated by 4Gy X-ray and the expression of MRE11 protein changes with Oh,1.5h,4h,6h, 8h,12h,24h.Method:Our studies were stedied with CNE1 cells cultured in vitro. The expression of MRE11mRNA in CNE1 cells was analyzed using RT-PCR after exposure to 0Gy,2Gy,4Gy,6Gy and 8Gy of ionizing radiation. The expression of MRE11 protein in CNE1 cells was analyzed using immuno-fluorescence assay before and after 4Gy X-ray exposure at different time points, such as 0h,1.5h,4h,6h,8h,12h,24h.Result:The semi-quantitative OD value of MRE11mRNA were 1.1±0.43,0.87±0.42,0.81±0.32,0.87±0.48,0.83±0.21 after OGy,2Gy,4Gy, 6Gy,8Gy irradiation, respectively. The One-way analysis of variance (Kruskal-Wallis test) shows P= 0.9797 (P> 0.05), the expression of MRE11 mRNA had not statistically significant between each group. We used immunofluorescence assay to detect MRE11 protein of CNE1 cells after irradiation by 4Gy at 0h,1.5h,4h,6h,8h,12h,24h time points. Fluorescence intensity first increased and then decreased. The strongest point of fluorescence intensity was 4-hour time point. The mean OD value of MRE11 protein were 138.7±24.91,238.1±19.57,721.5±14.60, 326.4±22.74,262.0±27.68,228.6±20.32,132.2±24.73.1.5h,4h,6h,8h,12h groups compared with the Oh group. Datum by One-way analysis of variance (Kruskal-Wallis test) shows P<0.05, the expression of MRE11 mRNA had statistically significant.24h group compared with the Oh group, P> 0.05, the difference was not statistically significant.Conclusions:The expression of MRE11 mRNA in CNEl cell lines was independent of X-ray dose escalation. But MRE11 protein expression with time first increased and then decreased, and 4h time point after X-radiation expressed the strongest.