人肺鳞癌组织的血清蛋白质组学的比较分析

采用以肿瘤免疫学与蛋白质组学(proteomics)研究技术有机地结合为基础的血清蛋白质组学研究体系(serologicproteomeanalysis ,SERPA)筛选肺癌分子标志物.对10例人肺鳞癌组织,应用双向凝胶电泳(two dimensionalelectrophoresis ,2 DE)技术对同一肺鳞癌组织的细胞总蛋白同时进行电泳后获得3张相同的凝胶,其中一块2 DE凝胶经银染显色作为平行胶,其余两块2 DE凝胶经电转膜将凝胶中的蛋白质转至硝酸纤维素(NC)膜上,然后分别与肺癌患者的自身血清以及正常对照血清进行Western印迹分析,获取Western印迹反应图谱.经计算机图像分析识别差异反应的蛋白质,然后与平行胶比较找出相应的差异反应蛋白质点.获得了分辨率较高的人肺鳞癌组织与患者的自身血清以及正常对照血清的Western印迹反应图谱;图像分析共识别36±8个差异反应的蛋白质;在平行胶上找到了匹配的差异反应蛋白质点.对2 0个差异蛋白质点进行了肽质指纹图分析,鉴定出14个与细胞生长增殖、细胞代谢、细胞周期调控、信号转导等有关的肺鳞癌相关抗原.通过血清蛋白质组技术对肺鳞癌组织进行的研究,建立了分辨率较高的人肺鳞癌组织与患者的自身血清以及正常血清的Western印迹反应图谱,成功鉴定14个肺鳞癌相关抗原,为进一步筛选用于肺鳞癌诊断、治疗和预后评估

Comparative Analysis of Serum Proteomes for Discovery of Biomarkers for Human Lung Squamous Carcinoma

Carcinogenesis of lung squamous carcinoma(LSC) is a complex process involving multiple events and steps. However,the definite diagnostic and prognostic protein biomarker for LSC are still unclear. To identify tumor-associated antigens by using serologic proteome analysis (SERPA), an approach which combines conventional proteome analysis with serological screening.SERPA of ten human lung squamous carcinoma tissues were performed. Three gels were run simultaneously with equal amounts of proteins from the same lung squamous carcinoma by IPG-2D PAGE. After separation, the proteins of one replica gel was visualized by silver-based staining technique. The other two gels were used for Western blot analysis. Proteins are transferred to NC membranes, then reacted with autologous patient serum and with control normal serum, respectively. The Western blot imaging films were obtained and the differential reacting protein spots were recognized, then the differential reacting protein spots in the replica gel by matching analysis. Well-resolved, reproducible 2-DE Western blot imaging films of human lung squamous carcinoma reacted with autologous patient sera and with control sera were obtained. 36 ±8 differentially expressed proteins which only were reactive with lung squamous carcinoma patient sera were found. Fourteen proteins, which are lung squamous carcinoma-associated antigens, were identified by peptide mass fingerpriting analysis with relative software. some proteins were the products of oncogenes, and others involve in the regulation of cell cycle and signal transduction. Well-resolved, reproducible 2-DE Western blot imaging films of human lung squamous carcinoma reacted with autologous patient sera and with control sera were obtained and fourteen tumor-associated antigens were characterized by using SERPA. These results may provide scientific foundation for screening the molecular biomarker used to diagnose and treat lung squamous carcinoma, as well as to elevate the patient's prognosis and provide new clue for the research of lung squamous carcinogenic mechanism.