应用二维电泳和质谱技术筛选食管癌及癌旁组织的差异表达蛋白质

目的 筛选食管癌组织与正常食管组织中的差异表达蛋白质,以发现用于食管癌早期诊断的生物学标志物.方法 收集食管癌组织及其配对的正常食管组织,提取组织蛋白质,进行二维电泳,选取在癌组织中高表达的3个点和在正常食管组织中高表达的3个点进行基质辅助激光解吸离子化-飞行时间质谱分析,将获得的肽质量指纹图进行生物信息学分析,鉴定差异蛋白质,利用RT-PCR方法验证蛋白质组筛选结果.结果 建立了分辨率高、重复性较好的食管癌和食管正常组织蛋白质的二维电泳图谱,经鉴定发现了鳞状细胞癌抗原1、细胞角蛋白4和膜联蛋白Ⅰ在食管癌组织中表达下调.磷酸丙糖异构酶1、锰超氧化物歧化酶和热休克蛋白27在食管癌组织中表达上调.RT-PCR的实验结果验证了差异蛋白质的可靠性.结论 食管癌组织和正常食管组织的差异表达蛋白质可做为食管癌早期诊断的候选生物学标志物.

Screening of differentially expressed proteins from human esophageal cancer and esophageal tissues by two-dimensional difference gel electrophoresis and mass spectrometry

objective To establish two-dimensional electrophoresis profiles from human esophageal cancer tissue and paired normal esophageal tissue and identify differentially expressed proteins to identifythe molecular markers for early-stage diagnosis. Methods The total proteins of human esophageal cancer tissue and paird normal esophageal tissue were separated by immobilized pH gradient-based two-dimensional gel electrophoresis (2-DE). The 6 differentially expressed proteins were analyzed by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS). The peptide mass fingerprintings (PMF) were identified by database searching. Six differentially expressed proteins were validated by RT-PCR. Results The well-resolved, reproducible 2-DE patterns of esophageal cancer tissue and esophageal normal tissue were established. Using MALDI-TOF-MS technology, 6 differential protein spots were identified. Among them, the expressions of squamous cell carcinoma antigen 1 (SCCA1b), KRT4 and annexin A1 were downregulated and triosephosphate isomerase (TPI1), heat shock protein 27 (HSP27) and manganese superoxide dismutase (MnSOD) were upregulated in esophageal cancer tissues. Conclusion The identification of differentiall exressed proteinsin human esophageal cancer and normal tissue will be helpful for screening the biomarker for early-stage diagnosis.