应用iTRAQ技术筛选DNT与低级别胶质瘤差异表达蛋白

目的 应用同位素标记相对和绝对定量蛋白质组学技术(iTRAQ)联合液相串联质谱筛选胚胎发育不良性神经上皮肿瘤与低级别胶质瘤的差异表达蛋白。 方法 收集胚胎发育不良性神经上皮肿瘤(编号113)与低级别胶质瘤(编号114)各6例实体组织冻存新鲜标本，各组标本混合，通过蛋白质提取，蛋白质浓度测量(采用Bradford定量)，聚丙烯酰胺凝胶电泳，蛋白质酶解，iTRAQ标记，SCX分离，再进行基于QE的液质联用分析，得到信息数据。使用蛋白质鉴定软件Mascot 2.3.02，选择UniProt-Human数据库，然后进行数据库搜索和生物信息学分析。依据蛋白质丰度水平，当差异倍数达到1.3倍以上，且经统计检验其P＜0.05时，视为差异蛋白。 结果 鉴定出了中国大陆黄种人DNT相对于低级别胶质瘤的差异蛋白质88个，其中上调蛋白质44个，下调蛋白质44个。这些差异蛋白具有不同生物学活性，并参与多种代谢及信号通路。其中重要的蛋白有水通道膜内在蛋白1、丝氨酸／苏氨酸激酶、细胞内氯离子通道蛋白1、膜联蛋白A1、谷氨酰胺合成酶、硫酸软骨素多糖蛋白4、S100A9、S100A16、S100A13、异柠檬酸脱氢酶等。 结论 iTRAQ技术实用可靠，有效筛选出胚胎发育不良性神经上皮肿瘤与低级别胶质瘤的差异表达蛋白。 

iTRAQ technology for differentially expressed proteins screening in dysembryoplastic neuroepithelial tumor and low grade glioma

Objective To screen differentially expressed proteins in dysembryoplastic neuroepithelial tumor(DNT) and low grade glioma(LGG) by the proteomics analysis using isobaric tags for relative and absolute quantification(iTRAQ) combined with liquid chromatography-tandem mass spectrometry(LC-MS/MS). Methods The fresh frozen solid tissues of DNT(6 cases) and LGG(6 cases) were sampled and mixed.Then the protein extraction from admix specimens,protein concentration measurement (Bradford),SDS electrophoresis,protein enzymolysis,iTRAQ mark,SCX,2D and LC-MS/MS were performed to get the data.The MS/MS data were searched against the International UniProt-Human using the Mascot 2.3.02 for peptide identification and quantification.According to protein abundance,differentially expressed proteins were determined when difference was more than 1.3 times and P<0.05. Results Total 88 differentially expressed proteins were identified between DNT and LGG in China Yellow.44 proteins were significantly up-regulated(>1.3-fold) and also 44 were significantly down-regulated.These proteins had different of biologic activity and participated many metabolisms and signal pathways.The important proteins include aquaporin 1,serine/threonine-protein kinase,chloride intracellular channel 1,annexin A1,glutamine synthetase,chondroitin sulfate proteoglycan 4,S100A9,S100A16,S100A13,isocitrate dehydrogenase and so on. Conclusion The differentially expressed proteins of DNT and LGG identified by proteomic analysis using iTRAQ are reliable.The iTRAQ technology can provide a good platform to identify moresignificant molecule difference biomarkers of DNT and LGG.