凋亡抑制蛋白c-FLIP高表达与白血病细胞耐药的关系  

Relationship between High Expression of c-FLIP and Drug Resistance of Leukemia Cells

作　　者：刘佳 王颖 李赛赛 徐颖茜 邢海燕[1] 唐克晶[1] 田征[1] 饶青[1] 王敏[1] 王建祥[1] 

机构地区：[1]中国医学科学院北京协和医学院血液学研究所实验学血液学国家重点实验室,天津300020

出　　处：《中国实验血液学杂志》2017年第6期1621-1626,共6页Journal of Experimental Hematology

基　　金：国家自然科学基金项目(81570147);中国医学科学院医学与健康科技创新工程(2016-I2M-1-001);天津市临床医学研究中心建设项目(15ZXLCSY00010)

摘　　要：目的:探讨抗凋亡蛋白c-FLIP的表达对于白血病细胞系Kasumi-1细胞耐药的作用及其机制。方法:应用Tet-on系统,通过强力霉素(doxycycline,Dox)调控c-FLIP的条件性表达;将c-FLIP基因与慢病毒目的载体p LVX-Tight-puro连接,感染Kasumi-1细胞并获得稳定表达c-FLIP蛋白的稳定克隆;应用实时荧光定量PCR和Western blot方法证实c-FLIP在mRNA和蛋白水平的过表达情况;通过流式细胞术分析Fas激动剂CH11和HDAC抑制剂苯丁酸钠处理的Kasumi-1细胞的凋亡变化,以及过表达c-FLIP蛋白后对凋亡发生的影响;应用吉姆萨染色观察细胞凋亡形态变化,Western blot检测caspase-8凋亡信号通路的激活情况。结果:CH11和苯丁酸钠能够诱导Kasumi-1细胞凋亡,在Kasumi-1细胞过表达c-FLIP蛋白后,细胞能够抵抗CH11和苯丁酸钠诱导的凋亡;Western blot显示c-FLIP过表达阻滞了caspase-8凋亡信号通路的激活。结论:c-FLIP基因高表达使白血病细胞系Kasumi-1细胞对CH11和苯丁酸钠产生耐药作用。Objective： To explore the effect of c-FLIP expression on drug resistance of Kasumi-1 leukemia cells and its mechanisms. Methods： Tet-on inducible system was used to construct the conditional expression vector of c-FLIP by cloning the c-FLIP gene into lentivirus vector p LVX-Tight-Puro,then the Kasumi-1 cells were transfected with lentivirus p LVX-Tight-Puro-c-FLIP. The expression of c-FLIP was induced by doxycycline（ Dox） for different time and doses,and verified by qRT-PCR and Western blot. On the basis of the overexpression of c-FLIP,the Kasumi-1-c-FLIP cells were treated with CH11 and PB in order to induce apoptosis,and the Giemsa staining was used to show the apoptotic cell morphology. Results： qRT-PCR and Western blot showed the overexpression of c-FLIP,the CH11 and PB can induce Kasumi-1 cell apoptosis,while the c-FLIP overexpression weakened this effects. Western blot showed that the c-FLIP blocked the caspase-8 activation. Conclusion： The overexpression of c-FLIP inhibits the apoptosis caused by CH11 and PB,and leads to drug resistance in leukemia cells.

关 键 词：C-FLIP 白血病 KASUMI-1细胞 白血病细胞耐药 

分 类 号：R733.7[医药卫生—肿瘤]