带状疱疹后疼痛治疗前后蛋白质组学生物标记特征研究

【作者】 陆景珊；

【导师】 张贵君；

【作者基本信息】 北京中医药大学 ， 中药学， 2006， 硕士

【摘要】 蛋白质组学是在后基因组时代出现的一个新兴的研究领域,它的主要任务是识别鉴定细胞、组织或机体的全部蛋白质。主要应用领域是医药市场和疾病的机理研究。带状疱疹后疼痛的确切机理目前尚不明确,蛋白质组学的发展使人们有了新的认识工具。本研究是治疗带状疱疹后疼痛中药“复元镇痛汤”研究的一部分,隶属于国家教育部重大项目——“中药复方功能主治与其药效物质基础的相关性研究”课题(课题编号00176)。1研究目的本研究通过荧光标记和双向电泳的方法考查带状疱疹后疼痛患者治疗前后CSF蛋白质表达的差异,探讨用蛋白质组学方法解释PHN发病机理的可行性,为最终能够找到该病的疾病特异性蛋白DSPs作准备,为中药的信息物质鉴定和药效组分生物筛选奠定基础。2研究内容和方法2.1 Ettan 2-D Quant定量试剂盒进行总蛋白含量测定;2.2分析型凝胶的样品CyDyeTM DIGE荧光标记。应用CyDyeTM DIGE荧光标记的双向凝胶电泳,分离并分析带状疱疹患者治疗前后CSF的蛋白质表达差异。通过荧光扫描及差异分析确定表达量差异显著的蛋白质点;2.3制备型凝胶电泳质谱分析,应用基质辅助激光解吸电离/飞行时间质谱(MALDI-TOF MS)确定这些蛋白质的种类。3研究结果所得荧光图像使用DeCyder差异分析软件处理比较。在凝胶电泳图谱2500个蛋白质点中,统计得出有效变化蛋白质点145个,其中浓度减少的蛋白质点103个,增加的42个。取其中的一个蛋白质点A,经酶解消化后,进行MALDI-TOF-MS分析,通过查询数据库得知该蛋白为Complement factor B。包括该蛋白在内,一共用质谱鉴定出32个蛋白质。4结论应用蛋白质组学的方法可以找到带状疱疹患者治疗前后CSF中差异表达的蛋白,并通过质谱和数据库查询确定差异表达蛋白的种类。证明该方法寻找带状疱疹的DSPs是可行的。为采用生物方法筛选带状疱疹后疼痛的药物及其药物的生物鉴定方法奠定了基础。5创新点5.1荧光染色的蛋白质组学方法在带状疱疹后疼痛机理研究中的应用。5.2蛋白质组学在中药信息物质鉴定领域的应用。更多还原

【Abstract】 Proteomics is a new research area in postgenome era. Its main task is to indentify all proteins in cells,tissues and organisms; its main application aera is medicine matket and mechanism research of diseases. Now a days, the mechanisms of postherpetic neuralgia (PHN) is not known and the developping proteomics gives us a new tool.This article is a part of the research of“Fu Yuan Zhen Tong Soup”, which belonged to a great subject of the National Ministry of Education----“the research in the relationship between the function and the drug action foundation of the Chinese medicine”(subject No.00176).1 ObjectiveThis research was designed to study the feasibility of explanation for mechanism of PHN by examing the differences of CSF protein expression of PHN patients after treatment and prepared for further research.2 Contents and methods2.1 Total protein level assay with Ettan 2-D Quant kit.2.2 The CyDyeTM DIGE fluorescent labelling for samples of analysis mode gel. Make use of two dimensional gel electrophoresis of CyDyeTM DIGE fluorescent labelling to determine the differences of protein expression between undergoing PHN patients and cured ones, and find out the differently expressed proteins by fluorescent scanning and distinction analysis.2.3 Mass chromatographic analysis for preparation mode gel. Determine protein species by the application of MALDI-TOF MS.3 ResultsAfter comparisons between photofluorograms by DeCyder analysis softwares, we got 2,500 pretein spots, among which 145 spots are effectively changed. 103 of the them are decreased and 42 are increased in concentration. We take one of the proteins named Protein A for example. After digestion,MALDI-TOF-MS analysis and searching database, we confirmed the name of Protein A as Complement factor B. Besides this protein, 32 proteins were identified.4 ConclusionsDifferently expressed proteins can be found out by using proteomics methods. After mass analysis and searching the database, we could find out the exact protein. The results of this paper demonstrate the feasibility of proteomics technogy in PHN studies and this paper also prepared for further research in DSPs of PHN. 5 New ideas5.1 The application of the fluorescent labelling in proteomics in the mechanism research in PHN.5.2 The application of proteomics in the identification of the information substances in Chinese medicine.更多还原