AnnexinⅤ-FITC/PI双标记流式细胞术检测结直肠癌细胞凋亡

Analysis of 5-FU-induced Apoptosis in Colo 320 Cell Line by Flow Cytometry Using Annexin V and Propidium Iodide Double Staining

目的:探讨化疗药物5氟尿嘧啶(5-FU)诱导肿瘤细胞凋亡的作用特性,同时分析AnnexinⅤ-FITC/PI双标记流式细胞术在检测细胞凋亡中的应用。方法:采用不同浓度的5-FU作用于培养的结直肠癌细胞株colo320,并设不同的时间段,应用流式细胞术和Hoechst33342荧光染色两种方法观察5-FU诱导肿瘤细胞凋亡的时间、剂量特性。结果:5-FU可以诱导colo320细胞的凋亡,80μg/ml作用4h后AnnenxinⅤ单阳性即开始增多,至12h,AnnenxinⅤ及PI双阳性即坏死细胞也开始增加。随药物浓度的提高,各组凋亡细胞所占比例也随之升高。流式细胞术与荧光染色的结果保持一致。结论:化疗药物5FU诱导肿瘤细胞凋亡作用呈时间、剂量依赖。AnnexinⅤ-FITC/PI双标记流式细胞术可用于检测早期的细胞凋亡事件,简单、灵敏,可作为检测细胞凋亡的首选方法。

Objective: To investigate the profile of 5-FU-induced apoptosis in tumor cell line and evaluate the usefulness of Annexin V and propidium iodide double staining for detecting apoptosis. Methods: Colon cancer cell line colo320 was incubated with 5-FU of different concentration for 2, 4,6, 8 and 12h respectively, apoptosis was measured using Hoechst33342 staining and a flow cytometry method that measures Annexin V and propidium iodide (PI) staining. Results: Apoptosis could be induced in colo 320 cell line by 5-FU,the percentage of AnnexinⅤ^+PI^-cells increased after being treated with 80ug/ml 5-FU for 4 h,both apoptotic and necrotic cell increased at 12h.Apoptotic cells were also identified by Hoechst33342 staining. Conclusion: Apoptosis induced by 5-Fu is time and dose dependent.Annexin V and PI double staining is a specific, sensitive, and quantitative method for analyzing apoptotic cells.