肺癌骨转移血清蛋白质组学研究

Serum proteomic study on lung cancer patients with bone metastases

目的:应用蛋白质组学双向荧光差异凝胶电泳和质谱技术筛选肺癌骨转移血清标志。方法:收集肺癌伴骨转移、无骨转移各24例血清样本。同组血清等量混合,用除白蛋白试剂盒除去血清中的白蛋白,再经除盐浓缩后,将4组处理后的血清样品用不同的CyDye染料交叉标记后进行双向荧光差异凝胶电泳(2-DDIGE)。利用DeCyder软件分析,将肺癌伴骨转移组与肺癌无骨转移组比较的差异蛋白质点行基质辅助激光解析离子化飞行时间质谱(MALDI-TOF-MS)鉴定。结果:双向荧光差异凝胶电泳显示,肺癌骨转移组血清蛋白中有16个斑点,与肺癌无骨转移组相比差异有统计学意义。其中11个蛋白质表达水平显著增加,选取5个蛋白质表达水平显著下降。5个差异蛋白点进行胶内原位酶解、肽质指纹图谱分析,成功得到4个蛋白质肽质指纹图,并查询数据库初步鉴定该4个蛋白质分别为视黄醇结合蛋白-甲状腺素运载蛋白复合物(RBP-TTR)、结合珠蛋白(Hp)、S-亚硝基血红蛋白(SNO-Hb)和de-sArg141αHb。结论:双向荧光差异凝胶电泳结合质谱鉴定是血清差异蛋白质组学研究的可靠平台和有力工具。所鉴定出的蛋白质可能与肺癌骨转移的发生、发展有关,有助于肺癌骨转移发病机制的了解。

OBJECTIVE：To screen serum biomarker candidates in lung cancer patients with bone metastasis using 2-D differential gel electrophoresis（2-D DIGE）and MS.METHODS：The serum samples from 24 lung cancer patients with bone metastases,24 lung cancer patients without bone metastases.The serum samples of the same group were pooled and the most abundant protein albumin was depleted.The concentrated and desalted samples were labeled with three different CyDyes including one internal standard,pooled from all the samples,and separated with 2-D DIGE in triplicate experiments.The biological variations of the protein expression level were analyzed with DeCyder software.The differentially expressed protein spots were analyzed by peptide-fingerprinting with matrix-assisted laser desorption/ionization time of flight mass spectrometry（MALDI-TOF-MS）.RESULTS：A total of 11 proteins were found to be significantly increased and 5 significantly decreased in the serum samples of lung cancer patients with bone metastases.The identified proteins included RBP-TTR complex that was decreased,and haptoglobin,SNO-Hb,desArg141 α Hb that were increased.CONCLUSIONS：The combination of 2-D DIGE and mass spectrometry strategy is an ideal platform and powerful approach for differential proteomic analysis of human sera.Four proteins might be involved in the process of lung cancer bone metastases and improve our understanding of the fundamental change that contributes to the process of carcinogenesis and the formation of metastases in lung cancer patients with bone metastases.