放射诱导HL-60细胞凋亡的流式细胞计量术分析

FLOW CYTOMETRY ANALYSIS OF RADIATION INDUCED APOPTOSIS IN HL 60 CELL LINE

目的 定性和定量地分析放射诱导ＨＬ６０ 细胞凋亡。 方法 以人类白血病细胞系ＨＬ６０ 为材料，应用光学显微镜和电子显微镜及琼脂糖电泳定性分析６０ Ｃｏ 放射线诱导ＨＬ６０ 细胞凋亡的发生，应用ＤＮＡ流式细胞计量术和末端转移酶（ＴｄＴ） 介导的缺口末端标记法（ＴＵＮＥＬ） 的流式细胞定量检测ＨＬ６０ 细胞凋亡。 结果 （１） 放射后ＨＬ６０ 出现细胞体积缩小，染色质固缩致密化、边缘化，染色体断裂，凋亡小体和细胞膜保持完整等形态学特征。（２） 琼脂糖电泳表现为特征性ＤＮＡ“梯谱”。（３）流式细胞计量术发现放射诱导ＨＬ６０ 细胞凋亡在照射后６ｈ 已较明显，而且细胞凋亡百分比随着照射剂量增加而增加。ＴＵＮＥＬ法检测细胞凋亡百分比值比ＤＮＡ流式细胞分析的方法得到的值要小，但差异无显著意义。 结论 流式细胞计量术结合细胞形态学检查和（ 或） 琼脂糖电泳能较好、简便、迅速地分析放射诱导ＨＬ６０

PURPOSE To analyze qualitatitively and quanititatively radiation induced HL 60 apoptosis. METHODS Apoptosis was induced in HL 60 cell line by irradiation. Light microscope,electron microscope, and agarose gel electrophoresis were performed to analyze apoptosis qualitatively. Flow cytometry was used to detect apoptotic cell DNA content and 3′ OH terminal of DNA fragment labeled by FITC dUTP.RESULTS (1)The irradiated HL 60 cell exhibited condensation of the cytoplasm,chromatin condensation,chromosal clumping and margination,and apoptotic body.(2)The apoptotic cell nuclear DNA was degradated with a characteristic “DNA ladder” subjected to electrophores.(3)Radiation induced HL 60 apoptosis showed a time and dose dependented curve with most apparent apoptosis at 6h after irradiation. The induced apoptotic cell percent measured by flow cytometry increased with dose.CONCLUSIONS Flow cytometry is a rapid,simple,sensitive,and specific method to analyze radiation induced apoptosis,however,its cell death must be confirmed by characteristic cell morphology or“DNA ladder”.