细胞分裂周期素25A影响放射线照射后HEp-2细胞的G_2/M期阻滞及凋亡

目的:研究放射线照射后HEp-2细胞的G2/M期阻滞与细胞分裂周期素(CDC)25A表达的相关性及过表达CDC25A蛋白对细胞周期及凋亡的影响。方法:通过流式细胞仪和Western blot方法检测4Gy X线照射后在不同时间点细胞周期及CDC25A蛋白含量;经脂质体转染pEGFP-N1-CDC25A质粒的HEp-2细胞为CDC25A组,转染pEGFP-N1质粒的为空载对照组,通过实时荧光定量PCR法检测这两组细胞中CDC25A的mRNA含量以鉴定转染效率;通过流式细胞仪检测CDC25A组和空载对照组细胞经4Gy放射线照射后细胞周期和细胞凋亡率;通过MTT法检测两组细胞的存活曲线。结果:放射线照射后G2/M期在细胞周期中的比例与CDC25A蛋白含量成正相关;CDC25A组细胞的CDC25A mRNA含量为空载对照组的15倍;过表达CDC25A联合4Gy放射线照射可废除G1/S关卡阻滞,促进G2/M期聚集和提前进入有丝分裂,可导致细胞凋亡增加和细胞存活率降低。结论:CDC25A的含量与G2/M期的比例正相关,调控CDC25A的表达可影响放射线照射后肿瘤细胞的细胞周期和凋亡水平。

CDC25A Influences the G2/M Phase Arrest and Apoptosis of HEp-2 Cells After Irradiation

Objective: To investigate the relationship between the G2/M arrest of HEp-2 after irradiation and CDC25A,and to study the effect of overexpressed CDC25A on the cell cycle and apoptosis.Methods: After irradiation with 4 Gy 6 Mev X ray,the cell cycle phase and CDC25A protein expression were determined by flow cytometry and Western blot respectively.Transfecting pEGFP-N1-CDC25A plasmid into HEp-2 cells(CDC25A group) caused CDC25A overexpression,but it was not found in control group cells(transfecting pEGFP-N1).The mRNA of CDC25A was analyzed by real time PCR in the two groups.After 4 Gy irradiation,the cell cycle and apoptosis of these two groups were determined by flow cytometry,and cell proliferation were assayed by MTT method.Results: After irradiation,the G2/M phase in the cell cycle was correlated with the level of CDC25A.We transfected the plasmid into cells successfully.The mRNA level of CDC25A in overexpression group was 15-fold more than that in control group;overexpression of CDC25A might abolish the G1/S arrest,and arrest in the G2/M phase,promote mitosis of HEp-2 cells,and decrease cell survival rate.Conclusion: CDC25A is correlated with G2/M ratio.With irradiation,regulating the expression of CDC25A could influence the cell cycle and apoptosis of HEp-2 cells.