慢性粒细胞性白血病干细胞抗伊马替尼机制的初步分析  

Primary study on the mechanism of chronic myelogenous leukemia stem cells against imatinib mesylate

作　　者：房佰俊[1] 宋永平[1] 林全德[1] 张艳莉[1] 魏旭东[1] 

机构地区：[1]河南省肿瘤医院,河南省血液病研究所,河南省郑州市450008

出　　处：《中国组织工程研究与临床康复》2007年第46期9263-9267,共5页Journal of Clinical Rehabilitative Tissue Engineering Research

基　　金：河南省杰出青年基金(0612000900);河南省医学科技创新人才工程项目(200590)~~

摘　　要：目的:了解伊马替尼对慢性粒细胞性白血病患者体内BCR/ABL融合基因阳性的原始及定向白血病干/祖细胞分化及增殖的影响,从而更深入阐明部分慢性粒细胞性白血病患者在经历一段血液学甚至是细胞遗传学水平上的完全缓解后复发及对伊马替尼耐药的机制。方法:实验于2006-09/2007-06在河南省血液病研究所完成。①对象:选取10例慢性粒细胞白血病患者,患者对治疗及实验知情同意。实验经医学伦理委员会批准。实验过程中应用的伊马替尼为Novartis公司产品,批号B-1701-0001-000005;氟波酯为淅川制药公司产品,批号030321495。②实验方法:抽取慢性粒细胞性白血病患者骨髓10mL,分离得到具有血液血管干细胞特性、BCR/ABL融合基因阳性的Flk1+CD31-CD34-细胞。以诱导造血集落的前48h,96h,120h内培养体系中含5μmol/L伊马替尼分别作为伊马替尼1,2,3组,设立空白对照,均体外培养18d,检测伊马替尼对造血集落培养基中的未分化及处于分化阶段的BCR/ABL融合基因阳性Flk1+CD31-CD34-细胞的增殖抑制作用。采用流式细胞术将BCR/ABL融合基因阳性Flk1+CD31-CD34-细胞分为3个亚群,即G0期、G1期及S/G2+M期,分别评估氟波酯、伊马替尼、氟波酯联合伊马替尼对3个亚群细胞的体外增殖及凋亡的影响。结果:①伊马替尼对处于分化阶段BCR/ABL融合基因阳性Flk1+CD31-CD34-细胞增殖及向血管内皮细胞分化的影响:与空白对照组比较,伊马替尼1组每1000个BCR/ABL融合基因阳性Flk1+CD31-CD34-细胞产生的粒-单系祖细胞集落数、红系爆式集落数无明显变化(P>0.05),伊马替尼2,3组两种造血集落数均显著降低(P<0.05)。在内皮诱导体系中与空白对照组比较,伊马替尼3组CD31+/vWF+细胞出现率明显降低(P<0.05),且分化形成血管样结构的时间延迟。②伊马替尼诱导BCR/ABL融合基因阳性Flk1+CD31-CD34-细胞及其分化细胞凋亡的比较:5μmol/L伊马替尼作用4ATM： To evaluate the effect of imatinib mesylate （STI571, Gleevec） on primitive/committed malignant progenitor cells in chronic myelogenous leukemia （CML） and to further elucidate the mechanisms involved in CML relapse and in some CML cells that are insensitive to STI571. METHODS： Experiments were performed at Henan Institute of Hematology from September 2006 to June 2007. ①Totally 10 patients with CML were selected. Informed consent was obtained from patients and Medical Ethics Committee in Henan Tumor Hospital before they enrolled. ST1571 （No. B-1701-0001-000005） was purchased from Novartis Company and 12-0-tetradecanoylphorbol-13-acetate （TPA） （No. 030321495） was purchased from Xichuan Pharmaceutical Company. ② About 10 mL bone marrow from each CML patients were obtained and BCR/ABL^＋FIk1^＋CD31^- CD34^- cells with hemangioblastic characteristics were isolated. The BCR/ABL^＋FIk1^＋CD31^-CD34^- cells cultured in Methocult GF＋ media with ST1571 for 48 hours, 96 hours or 120 hours respectively were acted as STI571 groups 1, 2 and 3 before they were induced to differentiate into hematopoietic colonies, while BCR/ABL^＋FIk1＋CD31^-CD34 cells in Methocult GF＋ media without ST1571 were investigated as blank control. They were all cultured in vitro for 18 days and inhibitory effect of STI571 on proliferation of differentiated and differentiating BCR/ABL^＋FIk1^＋CD31^-CD34^- cells was investigated. Then, G0, G1 or S/G2＋M fractions of BCR/ABL^＋FIk1^＋CD31^-CD34^- cells were isolated by fluorescence-activated cell sorting （FACS） and the effect of TPA combined with STI571 or TPA alone on these fractions was also studied. RESULTS： ①lnhibitory effect of STI571 on proliferation of BCR/ABL^＋FIk1^-CD31^-CD34 cells differentiating into endothelial cells： The ability of every 1 000 BCR/ABL^＋FIk1^＋CD31^-CD34^- cells to produce CFU-GMs （colony-forming units granulocyte-macrophage） and BFU-Es （burst forming unit-er-ythroid） between blank control group and S

关 键 词：干细胞 融合基因 慢性粒细胞白皿病 伊马替尼 氟波酯 

分 类 号：R394.2[医药卫生—医学遗传学]