应用基因芯片分析曲古菌素A促人白血病细胞株Molt-4凋亡的机制  

Mechanism of Apoptosis Induced by Trichostatin A in Leaukemia Molt-4 Cells Analyzed by Microarray

作　　者：洪振亚[1] 易莉莎[1] 苗新宇[1] 卢运萍[1] 周剑锋[1] 刘文励[1] 

机构地区：[1]华中科技大学同济医学院附属同济医院血液内科,湖北武汉430030

出　　处：《癌症》2006年第8期946-953,共8页Chinese Journal of Cancer

基　　金：国家基础研究重点规划"973"项目(No.2002CB513100);国家自然科学基金项目(No.30371657)~~

摘　　要：背景与目的:研究表明组蛋白去乙酰化酶(histonedeacetylase,HDAC)抑制剂曲古菌素A(trichostatinA,TSA)能以较低浓度诱导多种肿瘤细胞凋亡,但具体机制不甚明了。本实验拟研究TSA诱导人白血病细胞株Molt-4凋亡的作用及其分子机制。方法:应用细胞培养、MTT、PI单标流式细胞术、DNAladder观察TSA诱导Molt-4细胞和健康人外周血单个核细胞(peripheralbloodmononuclearcell,PBMC)凋亡的作用,用基因芯片(microarray)、逆转录聚合酶链反应(reversetranscription-polymerasechainreaction,RT-PCR)和Westernblot等方法检测特定浓度TSA作用Molt-4细胞一定时间后基因表达的差异。结果:TSA能以时间、浓度依赖性方式诱导Molt-4细胞凋亡,相同浓度和时间范围内TSA对PBMC无明显的诱导凋亡作用。TSA处理9h,基因芯片分析发现Molt-4细胞中表达下调的基因有313个。下调基因编码产物包括信号转导分子、转录因子、酶、受体等,其生物学功能包括调节细胞生长、分化发育、凋亡等。其中STAT5A下调80.4%,MYC下调77.3%,ikaros下调83.1%,半定量RT-PCR验证均显示TSA作用9h后这些基因的表达下调,同时STAT5A及MYC蛋白在TSA作用24h时表达也明显降低。结论:TSA能诱导Molt-4细胞凋亡并抑制其增殖,但对PBMC无明显作用。TSA对Molt-4细胞的这种选择性诱导凋亡和抑制增殖的机制与促进增殖、抗凋亡基因的变化有关。BACKGROUND ＆ OBJECTIVE： Histone deacetylase is overexpressed in a variety of cancers and is closely correlated with oncogenic factors. A histone-deacetylase inhibitor, trichostatin A （TSA）, has been shown to induce apoptosis in many cancer cells at submicromolar concentrations. However, the mechanism remains unknown. This study was to investigate the underlying mechanism of trichostatin A on apoptosis of Molt-4 cells by characterizing the global gene expression profiles before and after TSA treatment. METHODS： PI single-labeled flow cytometry, MTT and DNA ladder were used to observe the effect of TSA on apoptosis of MOLT-4 cells and normal human peripheral blood mononuclear cells （PBMC）. Microarray and reverse transcription-polymerase chain reaction （RT-PCR） and Western blot were used to detect the differentially expressed genes of Molt-4 cells after incubation with TSA. RESULTS, TSA could induce apoptosis in Molt-4 cells in a dose and time-dependent manner. Besides, the dose of TSA within the time duration which could induce significant apoptosis in Molt-4 cells did not demonstrate apparent cytotoxicity to PBMCs. After incubation with TSA for 9 hours, 313 genes were detected down-regulated by microarray. Proteins encoded by these genes included signal transduction molecules, transcription factors, enzymes etc., which were involeved in the regulation of cell growth, differentiation and survival. STAT5A, MYC and ikaros were down-regulated by 80.4%, 77.3% and 83.1%, respectively. The changes of the three genes were confirmed by RT-PCR and the changes of STAT5A and MYC were further confirmed by Western blot. CONCLUSION. The inhibition of cell growth and induction of apoptosis by TSA in Molt-4 cells may be due to the changes of pro-proliferation genes and anti-apoptosis genes.

关 键 词：曲古菌素A MOLT-4细胞 凋亡 基因芯片 基因差异表达 

分 类 号：R733.7[医药卫生—肿瘤]