中医不同证型帕金森病患者外周血单个核细胞蛋白质组学研究

Human Peripheral Blood Mononuclear Cell Proteome Analysis of Parkinson Disease with Different TCM Syndromes

目的研究中医肝阳化风证和血虚生风证帕金森病患者和正常人外周血单个核细胞(PBMC)的比较蛋白质组差异。从蛋白表达水平上探讨中医肝阳化风证和血虚生风证的本质。方法用固相pH梯度双向凝胶电泳分离肝阳化风证和血虚生风证帕金森病患者和正常人PBMC总蛋白质,考马斯亮蓝染色,PDQuest 2-DE软件分析,对部分差异蛋白质点进行基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)测定其胶内酶解后的肽质指纹图谱,用Mascot查询系统查询SWISS-PROT数据库。结果获得了分辨率和重复性均较好的双向电泳考染图谱,对其中的15个差异蛋白质点分别进行肽质指纹分析,经数据库查询,初步鉴定为一些与细胞骨架、抗氧化应激、蛋白降解、信号转导、细胞周期调控等有关的蛋白质。结论建立了帕金森病肝阳化风证和血虚生风证PBMC的双向凝胶电泳图谱,提示帕金森病肝阳化风证和血虚生风证患者和正常人的PBMC的蛋白质表达具有差异,这种蛋白质组的差异分析有助于为研究肝阳化风证和血虚生风证本质在蛋白质水平上提供物质基础。

Objective, To investigate the comparative proteomes difference of PBMC between health adults and Parkinson Disease （PD） patients with Ganyang Huafeng syndrome and Xuexu Shengfeng syndrome. So to approach the essence of the two TCM syndromes from the protein expression level. Methods A series of methods, including imrnobilolized pH gradient two-dimensional polyacrylamide gel electrophpresis （2-DE）, Coomassie Brilliant Blue staining, PDQuest 2-DE analysis software, peptide massfingerprint based on matrix-assisted laser desorption/ionization time of flying mass spectrometry （MALDI-TOF-MS） and SWISS-PROT database searching, were used to separate and indentify the proteome of PBMC between health adults and PD patients with Ganyang Huafeng syndrome and Xuexu Shengfeng syndrome. Results The good 2-DE pattern including resolution and reproducibility was obtained. After Coomassie Brilliant Blue staining, 15 spots were incised and analyzed by MALDI-TOF-MS. The typic peptide masses were searched in the SWISS-PROT database by Mascot software. All proteins were preliminarily identified, which ralated to cytoskeleton, anti-oxidative stress, protein degradation, signal transduction and cell cycle, etc. Condusions The well-resolved, reproducible 2-DE patterns of Ganyang Huafeng syndrome and Xuexu Shengfeng syndrome PD were established and revealed the protein expression difference of PBMC between the two syndromes of PD patients. This research is helpful to provide substance evidence in investigation of the essence of Ganyang Huafeng syndrome and Xuexu Shengfeng syndrome on protein level.