拓扑异构酶Ⅰ抑制剂对耐阿霉素淋巴瘤细胞株杀伤和诱导凋亡作用的研究  

The study of killing and proapoptotic effects of topoisomerase Ⅰ inhibitor on L1210/ADM cells

作　　者：方今女[3] 曹阳[2] 高文斌[1] 郑真真[1] 马建增[1] 黄剑辉[1] 吕金燕[1] 王刚[1] 李晓新[1] 

机构地区：[1]大连大学附属中山医院肿瘤科,116001 [2]大连大学生物工程学院 [3]延边大学医学部病理及病理生理学教研室

出　　处：《中国医药》2011年第9期1045-1047,共3页China Medicine

基　　金：辽宁省教育厅高等学校科学研究项目（20040053）;辽宁省大连市医学卫生科学研究计划项目（2007-83）

摘　　要：目的 研究拓扑异构酶Ⅰ抑制剂对耐阿霉素淋巴瘤细胞株（L1210/ADM）的杀伤和诱导凋亡的作用.方法 采用噻唑蓝（MTT法）、钙依赖性磷脂结合蛋白（Annexin V）染色、梯状DNA电泳与细胞形态学观察等方法分析不同浓度（0.05、0.10、0.20、0.40μmol/L）拓扑异构酶Ⅰ抑制剂对L1210/ADM细胞的杀伤和凋亡作用,采用半胱氨酸天冬氨酸蛋白酶（Caspase）8、Caspase-3抑制剂IETD-fmk、DEVD-CHO分析拓扑异构酶Ⅰ抑制剂介导的靶细胞杀伤和凋亡作用与Caspase的关系.结果 0.10 μmol/L拓扑异构酶Ⅰ抑制剂对靶细胞株的杀伤作用,与对照组和0.05 μmol/L组之间差异均有统计学意义（均P＜0.05）.加用Caspase酶特异性抑制剂3组各时间点的靶细胞存活率均高于单用拓扑异构酶Ⅰ抑制剂组,差异均有统计学意义（均P＜0.05）.靶细胞株经拓扑异构酶Ⅰ抑制剂处理后,部分靶细胞株出现了细胞凋亡;加用Caspase酶特异性抑制剂组中,有部分细胞死亡,但无凋亡小体形成,死亡细胞均为锥虫蓝染色.经拓扑异构酶Ⅰ抑制剂处理的靶细胞株大部分细胞出现胞体解离,并彼此黏附成簇状的、锥虫蓝拒染的凋亡小体,靶细胞株DNA电泳表现为特征性梯状图谱;加入Caspase酶特异性抑制剂的各组可以看到阻断了梯状DNA的产生.结论 拓扑异构酶Ⅰ抑制剂对L1210/ADM细胞株具有较强的杀伤和诱导凋亡作用,其机制可能涉及Caspase-8、Caspase-3的有序性活化.Objective To observe the killing and proapoptotic effects of topoisomerase Ⅰ （topotecan） inhibitor on L1210/ADM cells. Methods To analyze the the killing and proapoptotic effects of topoisomerase Ⅰ on L1210/ADM cells with MTT method, Annexin V, DNA ladder electrophoresis and cell morphology analysis. The relation between the killing and proapoptotic effects and Caspase using Caspase-8, inhibitor IETD-fmk and DEVDCHOs was observed. Results In contrast with 0. 05 μ mol/L group, the killing and proapoptotic effects of 0.10 μmol/L topoisomerase inhibitor was statistically different. With 3 groups of Caspase enzyme inhibitor at each time point, survival rate of target topoisomerase group was higher than that of the single-use topoisomerase Ⅰ inhibitor group（P 〈0.05）. After treatment with inhibitors of topoisomerase Ⅰ , some target cells died. In Caspase enzyme inhibitor group, some cells died but no apoptotic body formation, dead cells were trypan blue stained. Majority of target cells ever incubated by the topoisomerase Ⅰ inhibitors got dissociated and tufted with each other. Apoptotic bodies formed without trypan blue staining. Target cells electrophoresis showed the characteristic of DNA ladder pattern in the group with Caspase specific inhibitor of the enzyme can block the formation of DNA ladder. Conclusion Topoisomerase Ⅰ inhibitor has significant killing and proapoptotic effects on L1210/ADM cell lines, which can depend on activation of Caspase-8 and Caspase-3.

关 键 词：白血病 淋巴细胞 DNA拓扑异构酶类 Ⅰ型 细胞株 细胞凋亡 

分 类 号：R979.1[医药卫生—药品]