胆管癌肿瘤标志物的蛋白质组学初步研究

Primary proteomic study on tumor markers of cholangiocarcinoma

目的:蛋白质组学分析被认为是一种寻找潜在生物标志物,尤其是肿瘤标志物的有效方法。二维电泳蛋白质组学分析是一种流行和行之有效的技术。使用二维微分凝胶电泳(2-D DIGE)的研究是为了解癌症病人和对照组不同的蛋白质表达,并从蛋白质在二维图像上分布模式发现潜在的肿瘤标志物。方法:选取5例胆管癌病人和5例对照组胆管标本作为实验样本。然后提取DIGE裂解的蛋白质。随后提取的蛋白质以3种不同CyDyes进行标记,并通过2-D DIGE分离。结果:2-D DIGE从肿瘤和对照组标本中分离出多种蛋白质,点状分布在长13 cm、pH值在3.0～10.0的固相pH梯度胶条上。经过DeCyder软件的生物变异分析模块分析,所有胆管癌病人有20处蛋白质点被发现显着升高,5处蛋白质点下调(配对t检验,P<0.05)。然后由MALDI-TOF在16处蛋白质点上鉴定出13种蛋白质。结论:本研究采用2-D DIGE的方法对照研究胆管癌蛋白质组学,取得了胆管癌的二维蛋白分布图。经质谱鉴定的13种差异表达蛋白可作为潜在的肿瘤标志物以备进一步研究。

Objective Proteomic analysis is a useful method to search potential biomarkers of tumors.Two-dimensional（2-D） electrophoresis is a popular and proven separation technique for proteome analysis.This study aims to compare the different protein expression of tissues from patients with and without cancer using a two-dimensional differential gel electrophoresis（2-D DIGE）,and to identify protein patterns present in 2-D maps searching for potential tumor biomarkers.Methods Bile duct samples were obtained from 5 patients with cholangiocarcinoma and 5 control patients without cancer.Proteins were extracted with DIGE lysis,labeled with 3 different CyDyes,and separated by 2-D DIGE.2-D maps were visualized after coomassie blue staining and analyzed with the Image Master 2-D software.Different expression proteins were further identified by mass spectrometry（MS）.Results A large number of protein spots were separated in 2-D DIGE from the tumor and control groups on pH3.0-10.0 IPG strips of 13 cm.Using the biological variation analysis module of the DeCyder software,20 spots were found to be significantly up-regulated and 5 spots down-regulated in all patients（P0.05）.Thirteen out of 16 protein spots then were identified by MALDI-TOF.Conclusions By using a 2-D DIGE approach for proteomic research of cholangiocarcinoma,we obtained a 2-D protein maps of cholangiocarcinoma.Thirteen different express spots were identified by MS,which can be further evaluated as potential tumor markers.