冷休克诱导肿瘤细胞凋亡的动力学特征

KINETIC CHARACTERISTICS OF APOPTOSIS INDUCED BY COLD SHOCK IN HUMAN BREAST CANCER CELLS

应用扫描电镜观察、末端DNA转移酶介导的脱氧UTP缺口末端标记技术 (TUNEL)和流式细胞术研究低温冷休克诱导人乳腺癌细胞系 (ZR -75 -30)细胞凋亡 ,并对其细胞凋亡的动力学特征测定、分析。结果显示 :-5℃、0℃、4℃冷休克均可诱导ZR -75 -30细胞凋亡 ,但细胞凋亡进程受实验细胞预培养时间 ;冷休克温度、时间 ;细胞冷休克后恢复培养时间的影响 ,这些影响因素具有明显的动力学特征。经过对冷休克诱导ZR -75 -30细胞凋亡的动力学特征分析、比较 ,优选出研究冷休克诱导肿瘤细胞凋亡的实验体系 :即细胞经37℃预培养36h→冷休克 ( -5℃24h,0℃36h,4℃48h)→细胞冷休克后于37℃恢复培养18 -24h→细胞凋亡检测→检测结果综合分析。应用该实验体系可使细胞凋亡率达50-60 % ,且重复性尚佳。优化体系为深入研究冷体克诱导细胞凋亡的信号传递及其分子机制提供实验模型。

Apoptosis induced by cold shock in human breast cancer cell (ZR-75-30) was identified with scanning electron microscope and in situ end-labelling (TUNEL) and the apoptotic kinetic characteristics was studied with flow cytometry. The results showed that the cells exposed to cold shock (-5℃,0℃,4℃) exhibited characteristics of apoptosis, including nuclear condensation and fragmentation, microvilli disappearance, cell membrane blebbing and apoptotic bodies formation. All of these proved that cold shock can induce apoptosis in tumor cells. The quantitative analysis of cold shock inducing apoptosis by flow cytometry revealed that apoptosis in ZR-75-30 cells is correlated with preculture time, cold shock temperature, duration and rewarming time, showing obvious kinetics charaeteristics. In addition, an experimental system for studying cold shock inducing apoptosis was selected: Preculture of cells at 37℃ for 36h→Cold shock of cells (-5℃for 24h, 0℃ for 36h,4℃ for 48h) →Rewarming post-cold shock cells at 37℃ for 18-24h→Detecting apoptosis. It is an optimum experimental model to study signal transmisson and molecular mechanism of apoptosis induced by cold shock.