灭活细胞周期检测点激酶增强乳腺癌细胞放疗敏感性

目的:乳腺癌是严重威胁妇女健康的重要疾病,放疗常常作为乳腺癌综合治疗的重要手段。研究表明,临床上产生放疗抵抗的主要原因是由于放疗激活细胞周期检测点信号传导通路引起细胞自我修复而逃避凋亡,因而目前通过抑制细胞周期检测点信号通路增强肿瘤放、化疗敏感性,已成为国际上抗肿瘤治疗的一个重要方向和研究热点。Chk1和Chk2是细胞周期检测点中最重要的丝氨酸/苏氨酸激酶,本研究通过反义封闭Chk1和/或Chk2基因,阻断细胞周期检测点信号传导通路,研究对乳腺癌MDA-MB-231细胞放疗后细胞周期和凋亡的影响,从而评价Chk1和Chk2基因作为肿瘤治疗靶点的有效性。方法:Western-Blot法检测转染Chk1和Chk2正、反义寡核苷酸后,细胞内Chk1和Chk2蛋白表达情况;流式细胞仪AnnexinV-PI法和SubG1法检测单转染或联合转染Chk1/2反义寡核苷酸对放疗后MDA-MB-231细胞凋亡和细胞周期的影响。结果:反义封闭Chk1或Chk2基因可解除G2/M期阻滞,增强放疗后凋亡敏感性,而同时反义封闭Chk1和Chk2基因具有协同作用。结论:阻断细胞周期检测点信号通路的关键激酶Chk1和Chk2可显著增强放疗后凋亡敏感性。

Abrogation of cell cycle checkpoint kinases increases the radiation sensitivity to breast carcinoma cells

Objective:To block cell cycle checkpoint pathway by antisense blocking of Chk1 and /or Chk2 gene to increase the radiation sensitivity of breast carcinoma cell line MDA-MB-231,further evaluate the validity of Chk1/2 as the target gene in cancer therapy.Methods:Chk1/2 sODN and AsODN alone or in combination were transfected into MDA-MB-231 cells,the exposed cells to irradiation at 24 h after the transfection,the Chk1/2 protein change was assayed by Western-blot and the cell cycles and apoptosis rates were detected by FCM using AnnexinV-PI and SubG1 method.Results:The irradiated apoptosis sensitivity was increased by antisense blocking of Chk1 or Chk2 gene in MDA-MB-231 cells,furthermore,the G2/M phase blocking phenomenon decreased and a synergic effect was observed when Chk1 and Chk2 AsODN were used in combination.Conclusion:Abrogating the key effector Chk1 and Chk2 gene could increase the apoptotic sensitivity to irradiation.