中华血液学杂志
中華血液學雜誌
중화혈액학잡지
Chinese Journal of Hematology
2009年
6期
363-367
,共5页
于晓林%刘永%陈春燕%王涓冬%孔德晓
于曉林%劉永%陳春燕%王涓鼕%孔德曉
우효림%류영%진춘연%왕연동%공덕효
K562细胞%抗药性,多药%基因芯片%高三尖杉酯碱%基因,BMX
K562細胞%抗藥性,多藥%基因芯片%高三尖杉酯堿%基因,BMX
K562세포%항약성,다약%기인심편%고삼첨삼지감%기인,BMX
K562,cells%Drug resistance,multiple%DNA microarray%Homoharringtonine%Gene,BMX
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Western blot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mdr1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AL 049309(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Western blot法检测得到了相同的结果.RT-PCR和Western blot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.
目的 探討高三尖杉酯堿(HHT)誘導的人白血病細胞耐藥相關分子.方法 在前期建立的HHT誘導的人白血病多藥耐藥細胞株K562/HHT的基礎上,採用基因芯片技術比較K562/HHT細胞、其親本K562細胞以及用耐藥逆轉劑米非司酮(RU486)作用後的K562/HHT(K562/HHT/RU486)細胞三者基因錶達譜的差異,選擇在這三種細胞中呈動態變化的骨髓細胞X染色體上酪氨痠激酶(BMX)基因用RT-PCR和Western blot法在轉錄和翻譯水平進行驗證,進而轉染BMX基因到K562和K562/HHT細胞,觀察BMX過錶達時這兩種細胞內柔紅黴素(DNR)含量的變化,確定BMX是否在K562/HHT細胞耐藥形成中髮揮作用.結果 耐藥細胞K562/HHT與其親本K562細胞相比,共有117箇基因錶達有顯著差異,其中57箇基因錶達明顯上調,60箇基因錶達明顯下調,耐藥細胞K562/HHT中多藥耐藥基因mdr1錶達明顯上調;K562/HHT/RU486細胞與K562/HHT細胞相比,13箇基因錶達明顯上調,37箇基因錶達明顯下調.這些差異錶達的基因涉及耐藥、細胞信號傳導、細胞分化、細胞增殖、轉錄調節以及離子轉運等.基因NM-001721(BMX)、NM-031459(SESN2)、NM-033642(FGF13)和AL 049309(SFRS12)在兩組芯片中的錶達均有顯著差異.其中BMX基因在K562/HHT細胞中的錶達與K562細胞相比,明顯上調,在K562/HHT/RU486細胞則較K562/HHT細胞減低.進一步用RT-PCR和Western blot法檢測得到瞭相同的結果.RT-PCR和Western blot證實BMX質粒轉染的K562及K562/HHT細胞BMX錶達均上調,流式細胞術檢測到K562及K562/HHT細胞內DNR含量熒光彊度分彆為79.28±4.04和29.84±2.67,均較轉染前熒光彊度158.52±8.08和58.58±6.53顯著減低.結論 BMX在高三尖杉酯堿誘導的人白血病多藥耐藥細胞株K562/HHT耐藥性產生中髮揮作用.
목적 탐토고삼첨삼지감(HHT)유도적인백혈병세포내약상관분자.방법 재전기건립적HHT유도적인백혈병다약내약세포주K562/HHT적기출상,채용기인심편기술비교K562/HHT세포、기친본K562세포이급용내약역전제미비사동(RU486)작용후적K562/HHT(K562/HHT/RU486)세포삼자기인표체보적차이,선택재저삼충세포중정동태변화적골수세포X염색체상락안산격매(BMX)기인용RT-PCR화Western blot법재전록화번역수평진행험증,진이전염BMX기인도K562화K562/HHT세포,관찰BMX과표체시저량충세포내유홍매소(DNR)함량적변화,학정BMX시부재K562/HHT세포내약형성중발휘작용.결과 내약세포K562/HHT여기친본K562세포상비,공유117개기인표체유현저차이,기중57개기인표체명현상조,60개기인표체명현하조,내약세포K562/HHT중다약내약기인mdr1표체명현상조;K562/HHT/RU486세포여K562/HHT세포상비,13개기인표체명현상조,37개기인표체명현하조.저사차이표체적기인섭급내약、세포신호전도、세포분화、세포증식、전록조절이급리자전운등.기인NM-001721(BMX)、NM-031459(SESN2)、NM-033642(FGF13)화AL 049309(SFRS12)재량조심편중적표체균유현저차이.기중BMX기인재K562/HHT세포중적표체여K562세포상비,명현상조,재K562/HHT/RU486세포칙교K562/HHT세포감저.진일보용RT-PCR화Western blot법검측득도료상동적결과.RT-PCR화Western blot증실BMX질립전염적K562급K562/HHT세포BMX표체균상조,류식세포술검측도K562급K562/HHT세포내DNR함량형광강도분별위79.28±4.04화29.84±2.67,균교전염전형광강도158.52±8.08화58.58±6.53현저감저.결론 BMX재고삼첨삼지감유도적인백혈병다약내약세포주K562/HHT내약성산생중발휘작용.
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, K562 and K562/HHT/RU486 (K562/HHT reversed by RU486) cells were detected by DNA microar-ray. 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, 57 of which were up-regulated and 60 down-regulated. The mdrl gene was significantly up-regu-lated. When compared with K562/HHT, 50 significantly differently expressed genes were screened out in the K562/HHT/RU486 cells, 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 pCl-neo-BMX was transfected into K562 and K562/HHT cells, DNR concen-tration 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 multi-drug resistance of K562/HHT cell line.
