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目的:应用比较蛋白质组学的方法分析丙型肝炎患者外周血单个核细胞蛋白质表达模式的变化及寻找丙型肝炎相关生物标记分子,进一步识别鉴定其差异表达蛋白质,分析其对丙型肝炎慢性化机制的意义。方法:应用固相化pH梯度双向凝胶电泳(2-DE)分离健康者(10)及HCV患者(28)PBMC的总蛋白质,凝胶银染显色后,PDQuest图像分析软件进行比较分析、识别差异表达的蛋白质,应用基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)获得差异蛋白点的肽质指纹图谱,通过SWISS-PROT数据库鉴定蛋白质。结果:得到两张2-DE图谱,HCV患者及健康者PBMC凝胶的蛋白质点数分别为625及614;初步筛选出HCV患者与健康者存在明显差异的12个蛋白点,经质谱分析,初步鉴定了10种蛋白质。这些差异蛋白质包括病毒蛋白、蛋白质合成与分解、三大代谢相关酶类、细胞结构相关蛋白质以及信号转导相关蛋白质。结论:应用2-DE及MALDI-TOF-MS方法建立了HCV患者PBMC双向凝胶电泳图谱,分离并初步鉴定了10种与HCV感染相关的差异表达的蛋白质,为研究HCV慢性化相关机制提供新的线索。
OBJECTIVE: To compare the changes of protein expression patterns of peripheral blood mononuclear cells in patients with hepatitis C and to search for hepatitis C-related biomarkers using comparative proteomics and to further identify and identify differentially expressed proteins and to analyze their effects on chronic hepatitis C The significance of the mechanism. Methods: The total protein of healthy PBMCs (PBMCs) from healthy individuals (10) and HCV patients (28) was separated by two-dimensional electrophoresis (2-DE) using immobilized pH gradient gel electrophoresis. After gel silver staining, PDQuest image analysis software was used for comparative analysis. The differentially expressed proteins were identified. Peptide fingerprinting of differentially expressed protein spots was performed using MALDI-TOF-MS and proteins were identified by the SWISS-PROT database. Results: Two 2-DE maps were obtained. The protein spots of PBMC gels of HCV patients and healthy individuals were 625 and 614, respectively. Twelve protein spots with significant differences between HCV patients and healthy subjects were screened out, and identified by mass spectrometry 10 kinds of protein. These differential proteins include viral proteins, protein synthesis and breakdown, three major metabolic enzymes, cell-structure related proteins, and signal transduction related proteins. CONCLUSION: Two-dimensional gel electrophoresis of PBMC from patients with HCV was established by 2-DE and MALDI-TOF-MS. Ten differentially expressed proteins related to HCV infection were isolated and initially identified, providing novel mechanisms for studying the mechanism of chronic HCV infection The clues.