高三尖杉酯碱诱导的白血病耐药细胞基因表达谱的变化  

Changes of gene expression profile in homoharringtonine-induced leukemia multi-drug resistant cell line K562/HHT

作　　者：于晓林[1] 刘永[1] 王涓冬[2] 孔德晓[2] 陈春燕[1] 

机构地区：[1]山东大学齐鲁医院血液科,济南250012 [2]山东大学第二医院血液科

出　　处：《中华血液学杂志》2009年第6期363-367,共5页Chinese Journal of Hematology

基　　金：基金项目：山东省科技攻关重点项目（2004GG2202107）;山东省中青年科学家科技奖励基金（2004BS02007）;第五批山东省卫生系统“1020”科技人才工程

摘　　要：目的探讨高三尖杉酯碱（HHT）诱导的人白血病细胞耐药相关分子。方法在前期建立的HHT诱导的人白血病多药耐药细胞株K562／HHT的基础上，采用基因芯片技术比较K562／HHT细胞、其亲本K562细胞以及用耐药逆转剂米非司酮（RU486）作用后的K562／HHT（K562／HHT／RU486）细胞三者基因表达谱的差异，选择在这三种细胞中呈动态变化的骨髓细胞X染色体上酪氨酸激酶（BMX）基因用RT—PCR和Westernblot法在转录和翻译水平进行验证，进而转染BMX基因到K562和K562／HHT细胞，观察BMX过表达时这两种细胞内柔红霉素（DNR）含量的变化，确定BMX是否在K562／HHT细胞耐药形成中发挥作用。结果耐药细胞K562／HHT与其亲本K562细胞相比，共有117个基因表达有显著差异，其中57个基因表达明显上调，60个基因表达明显下调，耐药细胞K562／HHT中多药耐药基因mdr-1表达明显上调；K562／HHT／RU486细胞与K562／HHT细胞相比，13个基因表达明显上调，37个基因表达明显下调。这些差异表达的基因涉及耐药、细胞信号传导、细胞分化、细胞增殖、转录调节以及离子转运等。基因NM-001721（BMX）、NM-031459（SESN2）、NM-033642（FGF13）和AL049309（SFRS12）在两组芯片中的表达均有显著差异。其中BMX基因在K562／HHT细胞中的表达与K562细胞相比，明显上调，在K562／HHT／RU486细胞则较K562／HHT细胞减低。进一步用RT—PCR和Westernblot法检测得到了相同的结果。RT—PCR和Westernblot证实BMX质粒转染的K562及K562／HHT细胞BMX表达均上调，流式细胞术检测到K562及K562／HHT细胞内DNR含量荧光强度分别为79．28±4．04和29．84±2．67，均较转染前荧光强度158．52±8．08和58．58±6．53显著减低。结论BMX在高三尖杉酯碱诱导的人白血病多药耐药细胞株K562／HHT耐药性产生中发挥作用。Objective To study the resistant related molecules of human leukemia drug resistant K562 cells（K562/HHT） induced by homoharringtonine （HHT）. Methods Gene expression profiles on K562/ HHT, I（562 and K562/HHT/RU486 （ K562/HHT reversed by RU486） cells were detected by DNA microarray. The bone marrow tyrosine kinase gene in chromosome X（BMX） which changed dynamically among the three cells was confirmed by RT-PCR and Western blot. Then, BMX was transfected into K562 and K562/ HHT cells, and the changes of daunorubicin （DNR） concentrantions in these two cells were observed for BMX overexpression. Results As compared with K562, there were changes in 117 gene expressions in K562/HHT, 57of which were up-regulated and 60 down-regulated. The mdr1 gene was significantly up-regulated. When compared with K562/HHT, 50 significantly differently expressed genes were screened out in the K562/HHT/RU486 ceils, of which up- and down-regulated genes were 13 and 37 respectively. These genes involved in drug resistance, cell signaling, cell differentiation, cell proliferation, transcription regulator, ion transport and so on. Four genes [ NM-001721 （ BMX）, NM-031459 （ SESN2 ）, NM-033642 （ FGF13 ） and AL- 049309（SFRS12） ] expressed significantly differently in the two group cells, BMX gene expression was higher in K562/HHT, than in K562, but lower than in K562/HHT/RU486 as confirmed by RT-PCR and Western blot. After the plasmid pCI-neo-BMX was transfected into K562 and K562/HHT cells, DNR concentration was significantly lower （79.28 ±4.04,29.84±2.67） than those before transfection （ 158.52±8.08, 58.58 ±6.53 ）. Conclusion BMX is associated with multirug resistance of K562/HHT cell line.

关 键 词：K562细胞 抗药性 多药 基因芯片 高三尖杉酯碱 基因 BMX 

分 类 号：R686[医药卫生—骨科学]