rhLIF与IL-24基因协同诱导HL-60细胞的凋亡  

Synergistic effect of rhLIF and IL-24 gene on apoptosis of HL-60 cells

作　　者：郁心[1] 叶震敏[1] 盛伟华[1] 杨吉成[1] 缪竞诚[1] 

机构地区：[1]苏州大学医学院细胞与分子生物学教研室,苏州215123

出　　处：《中国免疫学杂志》2007年第9期772-777,共6页Chinese Journal of Immunology

基　　金：国防科工委基础科研计划资助项目(K0102061501);苏州大学医学发展基金资助项目(EE134517)

摘　　要：目的:构建能稳定表达白血病抑制因子(LIF)的转基因细胞,并研究所表达的LIF与IL-24基因在诱导HL-60细胞凋亡方面的协同作用。方法:用真核表达质粒pcDNA3-LIF转染ECV304细胞,G418筛选阳性细胞,收集阳性细胞和培养上清,RT-PCR检测LIFmRNA的表达。同时在已转化重组腺病毒质粒的大肠杆菌中抽提质粒pAdEasy-1-pTrack-CMV-IL-24,用PacI酶切使重组腺病毒质粒线性化,脂质体转染QBI-293A细胞,收获Ad-IL-24腺病毒子。将重组病毒子(Ad-IL-24)感染HL-60细胞,同时加入含LIF的培养上清,并设对照组,RT-PCR检测IL-24 mRNA表达,光镜下观察HL-60细胞形态的变化,激光扫描共聚焦显微镜观察细胞凋亡改变,流式细胞仪检测细胞凋亡率,免疫细胞化学分析凋亡因子的改变。结果:成功构建能稳定表达LIF蛋白的转基因细胞;获得高滴度的重组腺病毒Ad-IL-24,用它感染HL-60细胞后,能检测到IL-24 mRNA的表达。各种检测方法表明LIF、IL-24基因都能抑制HL-60细胞生长、诱导凋亡,且两者具有协同作用。结论:LIF、IL-24基因都能抑制HL-60细胞生长,诱导凋亡,两者具有协同作用。Objective：To comtruct the cells which can express LIF protein steadily and study the effect of rhLIF and IL-24 on cells. Methods： ECV-304 cells were infected by eukaryotic expression plasmid pcDNA3-LIF, then the positive cells were selected by G418. The positive cells were collected and their culture supematant was stored for further use. Expression of LIF mRNA was detected by RT-PCR. The pAdeasy-l-pAdTrack-CMV-IL-24 was extracted from DHSa. The recombined adenovirus vector was lineared with PacI and transfected into 293 cells. The IL-24 recombined adenovirus （Ad-IL-24） was obtained and used to infect HL-50 cells. At the same time, the culture supematant containing rhLIF was added into the ~ cells which was identified as positive expression of IL-24 by RT- PCR. Synergistic effect of the cytokines on HL-60 cells was tested by LSCM, FCM and iwanunohistochemistry stain assay. Results： The cells expressing LIF protein steadily were constructed successfully, the high titer of the recombined adenovirus（Ad-IL-24） was obtained. Expression of IL-24 in infected HL-50 cells was identified by RT-PCR. The apoptosis of HL-60 cells induced by rhLIF and IL-24 was proved by LSCM , FCM and iwanunohistochemistry stain assay. They had synergistic effect. Conclusion：rhLIF and IL-24 can inhibit growth of HL-60 cells and induce apoptosis of the cells. They have synergistic effect.

关 键 词：LIF IL-24基因 HL-60细胞 凋亡 

分 类 号：Q786[生物学—分子生物学]