用2-DE和MALDI-TOF-MS筛选多发性骨髓瘤患者骨髓上清液中标志物  

Screening biomarkers in bone marrow supernatant of multiple myeloma by 2-DE and MALDI-TOF- MS

作　　者：李佳[1] 赵冠飞[2] 房亚哲[1] 翟玉华[1] 陈文明[3] 吴琳[4] 孙卫红[1] 刘金英[1] 王清涛[1] 

机构地区：[1]首都医科大学附属北京世纪坛医院血液科,100038 [2]首都医科大学附属北京朝阳医院检验科 [3]首都医科大学附属北京朝阳医院血液科 [4]中国科学院北京基因组研究所

出　　处：《中华检验医学杂志》2011年第11期968-974,共7页Chinese Journal of Laboratory Medicine

基　　金：国家自然科学基金资助项目（30872391）

摘　　要：目的运用2-DE和MALDI—TOF—MS研究MM患者骨髓上清液中差异表达蛋白，以寻找对MM发病机制研究和诊断或鉴别诊断的特异性蛋白质标志物。方法收集14例MM患者、5例其他血液系统恶性肿瘤患者及5名健康对照者骨髓上清液标本。除去标本中白蛋白和免疫球蛋白G（IgG）后，用2-DE分离3组骨髓上清液标本。用ImageMaster 2D platinum5．0图像分析软件分析比较3组2-DE凝胶图像，以差异倍数在3倍以上的蛋白点作为候选差异表达蛋白。然后，用MALDI—TOF—MS对重复性较好的候选差异表达蛋白做PMF和二级质谱鉴定。将得到的肽片段质量进行NCBInr数据库检索，以鉴定出相应的差异表达蛋白。结果用图像分析软件对3组骨髓上清液2-DE凝胶进行分析比较，从MM组与健康对照组中筛选出47个差异蛋白点，从MM组与疾病对照组中筛选出58个差异蛋白点。从MM组选取41个差异点进行质谱分析并鉴定，发现MM组与其他2组相比，5种高表达蛋白为免疫球蛋白J链、K轻链、入轻链、前病毒遗传的Gag多聚蛋白和与半抗原结合的含成熟氧（末）端的催化抗体；3种低表达蛋白为血红蛋白、结合珠蛋白（Hp2）、锌-α2-糖蛋白。这些差异蛋白部分与MM的临床表现相关，部分与MM的发生发展及临床治疗相关。结论应用2-DE和MALDI—TOF—MS技术从MM患者骨髓上清液标本中筛出与MM疾病相关的8种差异表达蛋白，为进一步研究MM发生发展机制，完善MM诊断、鉴别诊断指标提供了参考依据。Objective To analyze the differentially expressed proteins in bone marrow supetnatants of multiple myeloma patients by using 2-DE and MALDI-TOF-MS, and search for the special protein markers for studying the mechanism of the development and diagnosis or differential diagnosis of multiple myeloma. Methods The bone marrow supernatant samples of fourteen muhiple myeloma patients, five other hematologic malignancies and five normal controls were collected. After removing albumin and IgG, the proteins in the supernatants were separated by 2-DE. Three groups images were analyzed and compared by ]magemaster 2D platinum 5.0 analysis software. Differentially expressed proteins were selected if the protein spots intensity showed more than 3 fold increase or decrease among different groups. The identities of the differentially expressed proteins with good repeatibility were determined by PMF based on by MALDI-TOF- MS or MALDI-TOF-MS/MS and NCBInr database search. Results 2-DE maps of bone marrow supernatants of the three groups could be analyzed and compared by image analysis of software. Forty-seven and fifty-eight differentially expressed protein spots were detected in multiple myeloma samples compared with normal controls and other hematologic malignancies samples, respectively. Forty-one reproducible spots were analyzed and identified by mass spectrum. Compared with other hematologic malignancies and normal controls, five up-expressed proteins and three down-expressed proteins were identified in multiple myeloma samples. They includes immunoglobulin J chain K and k light chain, provirus ancestral Gag polyprotein, mature oxy-cope catalytic antibody with hapten for up-expressed proteins, and hemoglobin, haptoglobin Hp2, zinc-alPha-2.-glycoprotein for down-expressed proteins. These differentially expressed proteins reflect the features of multiple myeloma, and relate to the development, progression and therapy of multiple myeloma. Conclusions Eight differentially expressed proteins in bone marrow supernatants of multiple m

关 键 词：多发性骨髓瘤 电泳 凝胶 双向 光谱法 质量 基质辅助激光解吸电离 生物学标记 

分 类 号：R733[医药卫生—肿瘤]