iTRAQ技术鉴定百草枯诱导大鼠肺纤维化模型中差异表达蛋白的研究

发布时间：2019-01-28 07:57

【摘要】：背景肺纤维化(pulmonary fibrosis,PF)是目前在呼吸系统中死亡率非常高的一种疾病,诱导其发病的因素也很多,PF的主要临床特征有呼吸困难逐渐递增和肺间质纤维增生呈弥漫性。能够引起PF的发病因素有很多,由于缺乏有效的药物来治疗,所以其预后的效果并不十分理想。百草枯(paraquat,PQ)是目前应用比较广泛的一种除草剂,百草枯中毒之后最主要靶点之一就是肺部,人体接触之后极易在肺部聚积,之后引起的肺水肿、低氧以及肺部急性损伤都可能导致肺纤维化的发生,但其作用机制目前还不十分清楚。本研究以百草枯诱导的大鼠肺纤维化模型为研究对象,分析鉴定大鼠肺纤维化肺组织之间的差异表达蛋白,以期寻找与肺纤维化相关的关键蛋白,为探索阐明肺纤维化形成、发生发展机制奠定重要的理论和实验基础。目的用同位素标记相对与绝对定量(iTRAQ)技术筛选百草枯诱导的大鼠肺纤维化模型特异性表达蛋白,以期找到一些与肺纤维化相关的蛋白。方法将20只雄性SD大鼠(180-200g)分为两组:模型组:灌胃给予50mg/kg百草枯,对照组:灌胃给予50mg/kg生理盐水。在第28天时分别处死两组大鼠,通过HE染色的方法观察大鼠肺组织学的改变;应用iTRAQ标记联合LC-MS/MS质谱检测分析蛋白的表达量;利用NCBI数据库和Proteome Discoverer软件分析蛋白表达间的差异。结果HE染色表明实验组大鼠肺纤维化分级大于对照组;iTRAQ技术分析鉴定出268个差异表达蛋白,其中表达上调蛋白194个,表达下调蛋白74个。结论通过筛选百草枯诱导的大鼠肺纤维化模型特异性表达蛋白,表明iTRAQ标记联合LC-MS/MS质谱技术可以对表达具有差异的蛋白进行相关研究,并通过筛选的方法获得与肺纤维化的发生和发展有关联的蛋白质,这将对了解肺纤维化的发生发展过程产生十分重大的意义,并且还可能为肺纤维化的治疗或者预后提供理论基础。
[Abstract]:Background Pulmonary fibrosis (pulmonary fibrosis,PF) is a disease with high mortality in the respiratory system at present. The main clinical characteristics of PF are the gradual increase of dyspnea and the diffuse proliferation of pulmonary interstitial fiber. There are many factors that can cause PF, and the prognosis is not very good because of the lack of effective drugs to treat it. Paraquat (paraquat,PQ) is one of the most widely used herbicides. One of the main targets after paraquat poisoning is lung. Hypoxia and acute lung injury may lead to pulmonary fibrosis, but its mechanism is not clear. In this study, the rat pulmonary fibrosis model induced by paraquat was used to analyze and identify the differentially expressed proteins in the lung tissues of rats with pulmonary fibrosis, in order to find the key proteins related to pulmonary fibrosis and to elucidate the formation of pulmonary fibrosis. The mechanism of occurrence and development lays an important theoretical and experimental foundation. Objective to screen the specific expression proteins of paraquat induced pulmonary fibrosis model by using isotope labeling and absolute quantitative (iTRAQ) technique in order to find some proteins related to pulmonary fibrosis. Methods Twenty male SD rats (180-200g) were divided into two groups: the model group was treated with 50mg/kg paraquat and the control group with 50mg/kg saline. On the 28th day, the rats of the two groups were killed, the changes of lung histology were observed by HE staining, and the expression of protein was detected by iTRAQ labeling and LC-MS/MS mass spectrometry. The differences of protein expression were analyzed by NCBI database and Proteome Discoverer software. Results HE staining showed that the grade of pulmonary fibrosis in the experimental group was higher than that in the control group, and 268 differentially expressed proteins were identified by iTRAQ technique, including 194 up-regulated proteins and 74 down-regulated proteins. Conclusion by screening the specific expression protein of paraquat induced pulmonary fibrosis in rats, it is suggested that iTRAQ labeling combined with LC-MS/MS mass spectrometry can be used to study the expression of differentially expressed proteins. The protein associated with the occurrence and development of pulmonary fibrosis is obtained by screening method, which will be of great significance in understanding the process of occurrence and development of pulmonary fibrosis. It may also provide a theoretical basis for the treatment or prognosis of pulmonary fibrosis.
【学位授予单位】：新乡医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R-332;R563

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