Ph阳性急性淋巴细胞白血病细胞株Sup-B15与基质细胞共培养后对伊马替尼敏感性的变化  

Effects of stromal cells on sentitivity to imatinib in Sup-B15 Philadelphia chromosome positive acute lymphoblastic leukemia cells

作　　者：张焕新[1] 陈翀[1] 闫志凌[1] 宋旭光[1] 陈伟[1] 李德鹏 邱婷婷[1] 张璞[1] 曾令宇[1] 李振宇[1] 徐开林[1] 

机构地区：[1]徐州医学院附属医院血液科,221000

出　　处：《中华血液学杂志》2015年第6期460-464,共5页Chinese Journal of Hematology

基　　金：国家自然科学基金（81300399）;江苏省高校自然科学基金（13KJD320003）

摘　　要：目的 探讨基质细胞OP9能否影响ph＋急性淋巴细胞白血病细胞株Sup-B15细胞对伊马替尼的敏感性以及作用机制.方法 实验分为Sup-B15细胞组及基质细胞OP9与Sup-B15共培养组（即Sup-B15/OP9组）.采用CCK-8法测定细胞增殖抑制率;流式细胞术检测Annexin V/7-AAD标记的细胞凋亡率、CD34＋CD38-细胞比例;荧光定量PCR检测细胞ALDH1、CD144、β-catenin mRNA水平;Western blot法检测细胞CD133、CD144、β-catenin蛋白表达水平;免疫共沉淀方法检测结合于CD144的β-catenin水平.结果 10～45 μmol/L伊马替尼对Sup-B15、Sup-B15/OP9细胞均有增殖抑制作用,其IC50值分别为26.3和35.8 μmol/L,两组差异有统计学意义（P＜0.05）.30 μmol/L伊马替尼作用于Sup-B15、Sup-B15/OP9细胞24h,细胞总凋亡率分别为（14.24±2.11）％和（3.45±0.68）％,两组差异有统计学意义（P＜0.05）.与Sup-B15组相比,Sup-B15/OP9组CD34＋CD38-细胞比例增高[（3.42±0.28）％对（2.16±0.15）％,P＜0.05],ALDH1 mRNA水平增高（0.097±0.012对0.046±0.010,P＜0.05）,CD133蛋白表达水平升高;CD144 mRNA水平增高（0.103±0.015对0.010±0.003,P＜0.05）,CD144蛋白表达水平亦明显增高;β-catenin mRNA水平无明显变化（P＞0.05）,其蛋白总量明显上升,转位到细胞核的β-catenin蛋白明显增加,结合于CD144的β-catenin蛋白增加.结论 与OP9细胞共培养后,Sup-B15细胞对伊马替尼敏感性下降,该作用可能与上调CD144表达、稳定CD144/β-catenin信号通路、增加β-catenin核转位有关.Objective To investigate the sensitivity of imatinib （IM） on Sup-B15 Ph＋ acute lmphoblastic leukemia （ALL） cells indused by stromal cells OP9,and to further explore its mechanism.Methods The study is divided into two group,Sup-B15 cells group and co-cultured with OP9 cells group （Sup-B15/OP9 group）.The inhibitory effects of IM on leukemia cells were measured by CCK-8 test,and the apoptosis by Annexin V/7-AAD dyeing and the percentage of CD 34＋CD38 leukemia cells were determined by flow cytometry.ALDH1,CD144,and β-catenin mRNA were detected by real-time RT-PCR,protein levels by Western blot.Inmunoprecipitation was used to detect the level of β-catenin connected to CD144.Results IM presented inhibitory effects on Sup-B15 and Sup-B15/OP9 cells at multiple concentrations from 10 μmol/L to 45 μmol/L.The IC50 of IM on Sup-B15/OP and Sup-B 15 cells were 35.8 μmol/L and 6.3 μmol/L,respectively （P＜0.05）.After 24 h of 30 μmol/L IM treatment,the percentages of apoptosis cells in Sup-B15/OP9 and Sup-B 15 cell were （14.24±2.11）％ and （3.45±0.68）％,respectively （P＜0.05）.The percentage of CD34-CD38-cells in Sup-B15/OP9 group was significantly higher than that in Sup-Bl5 group [（3.42±0.28）％ vs （0.16±0.15）％,P＜0.05].As compared to Sup-B15 cells,the transcription of ALDH1 in Sup-B15/OP9 group was remarkably upregulated （0.097±0.012 vs 0.046±0.010,P＜0.05）,and the CDl33 protein level was also upregulated in Sup-B15/OP9 group.The transcription of CD 144 in Sup-B15/OP9 group was remarkably upregulated compared with Sup-B 15 group （0.103 ± 0.015 vs 0.010±0.003,P＜0.05）,as well as the CD144 protein.β-catenin mRNA transcription has no obvious changes between Sup-B15 group and Sup-B15/OP9 group （P＞0.05）,while the whole β-catenin protein and the cell nucleus β-catenin significantly increased,as well as the β-catenin protein combined with CD144.Conclusions Co-cultured with OP9 cells,Sup-B15 cells show less sensitivity to imatinib.The raising activity of C

关 键 词：费城染色体 白血病 淋巴细胞 急性 抗原 CD144 Β-CATENIN 基质细胞 

分 类 号：R733.71[医药卫生—肿瘤]