肺结核患者外周血单个核细胞蛋白质差异的表达

目的 应用比较蛋白质组学方法筛选出肺结核患者发病免疫相关蛋白质,为阐明结核病的发病的免疫机制和诊断治疗提供理论依据.方法 收集10例肺结核患者和10名健康志愿者外周血单个核细胞(PBMC),采用双向凝胶电泳分离细胞总蛋白,运用Image Master 2D Elite 5.0图像分析软件识别差异表达的蛋白质点,应用基质辅助激光解析电离飞行时间质谱获取肽质量指纹图谱,检索数据库鉴定差异表达的蛋白质点,并对差异表达的蛋白质用Western blot进行验证.两组间比较采用秩和检验.结果 建立了重复性良好的肺结核患者和健康志愿者PBMC的双向凝胶电泳图谱,找出14个表达量有明显差异的蛋白质点并鉴定出共12种蛋白质,其中冠蛋白1A等10种蛋白在肺结核组中表达上调,普列克底物蛋白(PLEK)和Ras基因抑制蛋白1(RSU1)在肺结核组中表达下调;Western blot法验证PLEK在肺结核组表达下调,与双向凝胶电泳结果一致.结论 对肺结核患者PBMC的比较蛋白质组学研究鉴定出冠蛋白1A、PLEK等12种差异蛋白质,其中PLEK和冠蛋白1A可能通过对单核巨噬细胞吞噬作用的调节在肺结核发病中起着重要作用.

Differential expression of proteins in peripheral blood mononuclear cells from patients with pulmonary tuberculosis

Objettive To screen the differential expressed proteins of peripheral blood mononuclear cells(PBMC)in patients with pulmonary tuberculosis by comparative proteomics.Methods PBMCs from 10 patients with tuberculosis and 10 healthy volunteers were collected.The total proteins were extracted and separated by two-dimensional gel electrophoresis(2-DE).The sites of differential expression were analyzed using ImageMaster 2D Elite 5.0 image analysis software,and the peptide mass finger-printing(PMF) identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry(MalDI-TOF-MS).The biological information of these proteins was searched in the database and confirmed by Western blot.Results Well reproducible 2-DE maps of PBMC from the patients with tuberculosis and normal controls were established. Forteen differential protein spots were found.Twelve proteins were analyzed and identified.PIeckstrin and Ras suppressor proteinl were decreased in patients with tuberculosis while the other 10 proteins were increased.The decreased expression of Pleckstrin in tuberculosis Was confirmed by Western blot. Conclusions Analysis of differential protein expression in PBMC from patients with pulmonary tuberculosis by comparative proteomics identified 12 proteins.The results suggest that Pleckstrin and Coronin-1 A may have an important role in the pathogenesis of pulmonary tuberculosis through regulating the phagocytosis of mononuclear macrophages.