AO_EB染色法及流式细胞术检测DMF诱导人肝细胞凋亡.pdfVIP

浙江预防医学 2007 年第 19 卷第 6 期 　Zhejiang Prev Med , June 2007 , Vol 19 , No. 6 ·11 · AO/ EB 染色法及流式细胞术检测 DMF 诱导人肝细胞凋亡 1 2 1 路艳艳 　杨锦蓉 　王菁 　　【摘要】目的 　观察二甲基甲酰胺 (DMF) 诱导正常成人离体肝细胞凋亡情况。方法 　以 50mmol/ L 、 100mmol/ L 、150mmol/ L 和200mmol/ L 剂量DMF 对正常成人离体肝细胞染毒。采用AO/ EB 双重染色法进行形态 学观察 ; 流式细胞术检测细胞凋亡率变化。结果　AO/ EB 染色可见 : 活细胞核染色质着绿色 , 死亡细胞核染 色质着橘红色 , 晚期凋亡细胞核染色质为橘红色并呈固缩状或圆珠状 , 早期凋亡细胞核染色质着绿色呈固缩 状或圆珠状。染毒组早期凋亡细胞多于阴性对照组 , 随剂量增高 , 晚期凋亡细胞和死亡细胞也增多。不同剂 量组间染毒 24 小时后 , 流式细胞术散点图可见 , 随染毒剂量增加 , 活细胞率逐渐下降 , 以200mmol/ L 剂量组 的活细胞率最低。经单因素方差分析 , 空白对照组和各剂量组间早期凋亡细胞占细胞总数的百分率差异有统 计学意义 (F = 8299 , P 001) , 随染毒剂量增高 , 早期凋亡细胞率增加。结论 　一定浓度的 DMF 能诱导正 常成人离体肝细胞发生凋亡 , 其早期凋亡率有随 DMF 浓度的增高而增高的趋势。 【关键词】DMF 染毒 ; 肝细胞 ; 凋亡 ; AO/ EB 染色 ; 流式细胞术 中图分类号: R1351 　　文献标识码 : A 　　文章编号 : (2007) DMFInduced Apoptosis in Human Hepatic Cells Detected by Methods of AOEB Doubly Staining and Flow Cytometer LU Yanyan , YANG Jinrong , WANG Jing . ( Institute of Environmental Medicine of Zhejiang University , Hangzhou , Zhejiang , 310031 , China. ) 【Abstract 】Objective 　To analysis N , Ndimethylformamide (DMF) induced apoptosis in human hepatic cells. Methods 　 Human hepatic cells were exposed to DMF with doses of 50 mmolL , 100 mmolL , 150 mmolL and 200 mmolL , respectively. The morphology of hepatic cells was observed by acridine orangeethidium bromide (AOEB) double staining , and the rate of apoptosis was detected by flow cytometer ( FCM) . Results 　The result of AOEB double staining showed that the nuclear chromatins of living cells were green and those of dead cells were orange. The nuclear chromatins of nonviable apoptotic cells were orange which showed condensation , and that of viable apoptotic cell were green which showed condensation. There