摘要目的 寻找膀胱尿路上皮癌(BUC)患者与健康志愿者尿液蛋白质组学差异,为BUC的早期诊断奠定基础.方法 采用25％乙醇沉淀法制备尿液双向电泳样品,尿液蛋白经双向凝胶电泳(2-DE)分离后行考马斯亮蓝染色,利用PDQuest8.0 2D凝胶图像分析软件进行分析,所得差异蛋白点用基质辅助激光解吸电离飞行时间串联质谱仪(MALDI-TOF-TOF-MS)进行肽序测定,结合Swiss-Prot蛋白质数据库对差异蛋白进行初步鉴定,并应用Western blot方法对差异蛋白在两组间的差异表达情况进行验证.结果 获得了分辨率和重复性均较好的双向电泳图谱.BUC组与对照组凝胶的蛋白质点数分别为(789±18)个和(762± 14)个；与对照组比较,BUC组有6个蛋白质点表达明显下调,11个蛋白质点表达明显上调.经质谱分析后,初步鉴定了5种蛋白质,分别为纤维蛋白原、乳酸脱氢酶B、载脂蛋白A1、丛生蛋白和触珠蛋白,均在BUC组呈表达增高趋势.Western blot验证了双向凝胶电泳结果.结论 BUC患者与健康志愿者的尿液蛋白质组存在明显的差异,差异蛋白的鉴定为进一步寻找BUC早期诊断的分子标志物提供了研究基础和依据.

abstractsObjective To investigate the difference in urinary proteome between patients with bladder urothelial carcinoma (BUC) and healthy volunteers and to provide a basis for the early diagnosis of BUC.Methods The urine samples from BUC patients and healthy volunteers (controls) were treated by 25％ethanol precipitation and two-dimensional gel electrophoresis (2-DE),and the obtained urinary proteins were subjected to Coomassie brilliant blue staining and analysis by PDQuest 8.0 (2-DE image analysis software); the differentially expressed proteins were sequenced by matrix-assisted laser desorption/ionization time-of-flight/time-of-flight mass spectrometry and identified using the Swiss-Prot database; the differential expression of these proteins was verified by western blot.Results High-resolution and high-reproducibility 2-DE images were obtained from the urine samples of BUC patients and controls,with 789 ±18 and 762 ±14 protein spots,respectively.Compared with the control group,the BUC grouPhad significantly decreased expression of 6 protein spots and significantly increased expression of 11 protein spots.The mass spectrometry revealed five proteins with increased expression in the BUC group,including fibrinogen,lactate dehydrogenase B,apolipoprotein A1,clusterin,and haptoglobin,and the results were confirmed by western blot.Conclusion There is significant difference in urinary proteome between BUC patients and healthy volunteers; the identification of differentially expressed proteins in urine lays the foundation for identifying potential molecular markers in early diagnosis of BUC.