PSMB5基因G322A突变及硼替佐米耐药的JurkatB细胞与其亲本细胞蛋白质表达谱的差异研究  

Comparison of Protein Expression Profiles between Bortezomib-resistant JurkatB Cells with PSMB5 Mutation and Their Parent Cells

作　　者：吕书晴[1] 杨建民[1] 黄崇媚[1] 许晓倩[1] 周虹[1] 宋宁霞[1] 王健民[1] 

机构地区：[1]第二军医大学长海医院血液内科,上海200433

出　　处：《中国实验血液学杂志》2011年第4期869-873,共5页Journal of Experimental Hematology

基　　金：上海市科学技术委员会基金(编号07ZR14146;064119511);国家自然科学基金(编号30873042)

摘　　要：本研究旨在探讨具有PSMB5基因G322A突变且对蛋白酶体抑制剂硼替佐米耐药的T淋巴母细胞淋巴瘤/白血病细胞系JurkatB与亲本Jurkat细胞的细胞生物学特性和蛋白质表达的差异,确认JurkatB5(硼替佐米筛选终浓度500 nmol/L)及JurkatB8(筛选终浓度800 nmol/L)细胞对硼替佐米的耐药性和对常规化疗药物的敏感性。采用台盼蓝拒染活细胞计数绘制生长曲线,应用软琼脂培养法观察细胞集落形成率,用流式细胞术行细胞周期分析,实时荧光定量RT-PCR检测多药耐药相关基因MDR1、LRP、MRPmRNA表达,采用含有720个蛋白抗体的SpringB io抗体芯片检测细胞株间蛋白质表达差异。结果表明,JurkatB5和JurkatB8细胞与亲本Jurkat细胞相比,对硼替佐米48小时耐药倍数分别为33.52,39.04。48小时化疗药物杀伤实验表明,JurkatB5和JurkatB8细胞对柔红霉素、阿霉素、长春新碱、依托泊甙均无交叉耐药,耐药细胞株与亲本Jurkat细胞的增殖、集落形成和细胞周期分布差异无统计学意义(p>0.05)。JurkatB5细胞与Jurkat细胞MDR1、LRP、MRP基因mRNA表达无显著差异(p>0.05)。蛋白芯片杂交信号扫描共有264个可分析表达点,252个蛋白在3个细胞株中表达无明显差异(2倍以内),其中包括与多药耐药相关的15个蛋白。在JurkatB5或JurkatB8中较Jurkat细胞表达升高或降低2倍以上的蛋白共12个(细胞分裂周期蛋白34、细胞分裂周期蛋白37、CD34 II型蛋白、基质金属蛋白酶-2、细胞黏合素、高尔基体复合物、内皮蛋白、组蛋白去乙酰化酶1、穿孔蛋白、促乳蛋白、维甲酸受体β、整合蛋白β1),但未发现在JurkatB5和JurkatB8中较Jurkat细胞中表达同时升高或降低2倍以上的蛋白。结论:具有PSMB5基因G322A突变且对硼替佐米耐药的细胞株JurkatB与亲本Jurkat细胞相比较其常规细胞生物学特性无显著改变,无多药耐药表型及多药耐药相关基因的过表达。突变细胞株中大部分已�This study was purposed to investigate the differences of cytobiological characteristics and protein expression levels between bortezomib-resistant T-lymphoblastic lymphoma/leukemia cell lines JurkatB containing PSMB5 G322A mutation and their parent cell line Jurkat, The cytotoxicities of bortezomib and chemotherapeutic drugs to Jur- katB5 cells （end selection concentration of bortezomib was 500 nmoVL）, JurkatB8 （end selection concentration 800 nmol/L） and Jurkat cells were analyzed. The cell growth curves were drawn with viable cell counts by trypan blue assay, the colony formation rate were assayed by soft-agar colony culture, and the cell distributions in cell cycle were analyzed by flow cytometry, mRNA expression levels of multidmg resistance （MDR） genes MDR1, LRP and MRP were measured by real-time fluorescence quantitative RT-PCR, the differences of protein expression levels were detected by SpringBio antibody microarray containing 720 proteins. The results showed that the drug resistance multiples for 48 hours of JurkatB5 and JurkatB8 cells （relative to Jurkat） to bortezomib were increased by 33.52 and 39.04 times, respectively.JurkatB5 and JurkatB8 cells did not display significant cross-resistance to daunorubicin, adriamycin, vindesine, and etopside after exposure for 48 hours. There were no significant differences in the cell growth curve, colony formation rate and cell distributions in cell cycle between JurkatB5, JurkatB8 and Jurkat cells （p 〉 0.05 ）. There were no significant differences of mRNA expression levels of MDR1, LRP, MRP between JurkatB5 and Jurkat cells （p 〉 0.05 ）. There were 264 analyzable expression points detected by antibody microarray. Among them, 252 protein expression levels were not significantly different between JurkatB5, JurkatB8 and Jurkat cells （ 〈 2-fold）, including 15 drug resistance-related proteins. 12 proteins were detected at higher or lower expression levels in JurkatB5 or JurkatB8 cells then that in Jurkat cells （ cell division cycle

关 键 词：PSMB5基因 G322A突变 硼替佐米 JURKAT细胞 JurkatB细胞 蛋白表达谱 

分 类 号：R733.1[医药卫生—肿瘤]