阿尔茨海默症生物标记物的蛋白质组学研究

发布时间：2018-06-19 21:13

本文选题：阿尔茨海默症 + 蛋白质组学　；参考：《深圳大学》2017年硕士论文

【摘要】：目的:阿尔茨海默症(Alzheimer’s disease,AD)是一种神经退行性疾病,病变过程较长,出现记忆障碍、生活不能自理等病状,早发现早干预十分重要。现今神经影像学可以直观的显示患者海马的受损程度,判断病变进程,但是,神经影像检测价格昂贵,且需要患者长时间静卧不动对海马进行检测,有一定难度。而生物标志物的兴起,提供了另一条诊断途径。在生物标志物研究方面,脑脊液检测被认为是“金标准”,但取样困难,而外周血可以在一定程度上反映病情,且取样容易,易于推广,使用外周血来寻找阿尔茨海默症的生物标志物,便于疾病的诊断。本研究以健康对照组、AD组和轻度认知功能障碍患者(mild cognitive impairment,MCI)组为研究对象,选择适合的蛋白质组学研究技术,基于将这些技术适用性和优缺点,组合运用于实验中,筛选出阿尔茨海默症的候选生物标志物,并通过多种手段进行验证,致力于反复验证以确保结果的准确性,从候选生物标志物中鉴定出可信的生物标志物。方法:本研究第一部分以AD和健康对照为研究对象,利用酶联免疫检测技术(enzyme linked immunosorbent assay,ELISA)验证既往文献报道的AD生物标志物,包括:Apolipoprotein A1、Haptoglobin、Alpha-1-antitrypsin、Clusterin、C-reative protein、Matrix metalloproteinase 9、Calgranulin A、Calgranulin B和Alpha-2-macroglobulin。第二部分以AD和健康对照为研究对象,运用同位素标记相对与绝对定量技术(isobaric tags for relative and absolute quantitation,iTRAQ)和蛋白芯片技术,结合生物信息学分析,研究阿尔茨海默症生物标志物,同时用ELISA和免疫印迹技术(Western blot,WB)对部分差异蛋白进行了验证,包括:Afamin、Platelet basic protein、Fibulin-1、Plasma protease C1inhibitor、Thrombospondin-1、Inter-alpha-trypsin inhibitor heavy chain H4、Ceruloplasmin和Fibrinogen gamma chain。第三部分以MCI和健康对照为研究对象,运用iTRAQ技术研究两组血液表达差异蛋白。结果:第一部分,验证了文献中的部分生物标记物,一共有6个蛋白的表达水平有统计学意义,其中Haptoglobin显著上调,Apolipoprotein A1、Alpha-1-antitrypsin、Matrix metalloproteinase 9、Calgranulin A和Calgranulin B显著下调,这6个蛋白中Apolipoprotein A1、Calgranulin A和Calgranulin B的趋势与文献报道一致。第二部分,验证了运用iTRAQ和蛋白芯片技术筛选出的7个蛋白,其中Platelet basic protein、Fibulin-1和Fibrinogen gamma chain显著上调,Plasma protease C1 inhibitor、Thrombospondin-1、Afamin和Ceruloplasmin显著下调,这7个蛋白中Plasma protease C1 inhibitor的趋势与文献报道和iTRAQ结果趋势一致,Fibulin-1和Thrombospondin-1的变化趋势与iTRAQ结果一致,Ceruloplasmin和Fibrinogen gamma chain的变化趋势与文献报道、蛋白芯片结果和WB结果一致。第三部分,以对照组和MCI组为实验对象,运用iTRAQ技术筛选出差异蛋白。结论:本研究结果表明阿尔茨海默症的病变可能与Apolipoprotein A1、Calgranulin A、Calgranulin B、Plasma protease C1 inhibitor、Thrombospondin-1、Ceruloplasmin和Fibrinogen gamma chain的表达水平变化有关,这些蛋白可能是阿尔茨海默症的潜在生物标记物。本研究发现的这些蛋白可否作为阿尔茨海默症的生物标记物,尚需进一步研究。
[Abstract]:Objective: Blzheimer (Alzheimer 's disease, AD) is a neurodegenerative disease with a longer pathological process, memory disorder, and inability to take care of itself. Early detection and early intervention is very important. Neuroimaging can directly display the damage degree of the hippocampus and judge the process of the disease. However, the price of neuroimaging is used to detect the price. It is difficult to detect the hippocampus for a long time, and it is difficult to detect the hippocampus for a long time. And the rise of the biomarker provides another way of diagnosis. In the study of biomarkers, the detection of cerebrospinal fluid is considered as a "gold standard", but the sampling is difficult, and the peripheral blood can reflect the condition to a certain extent, and the sampling is easy and easy to sample. To promote the use of peripheral blood to find the biomarkers of Alzheimer's disease and facilitate the diagnosis of disease. This study was conducted in the healthy control group, the AD group and the mild cognitive dysfunction (mild cognitive impairment, MCI) group, and selected suitable proteomics research techniques based on the applicability and advantages and disadvantages of these techniques. In the experiment, the candidate biomarkers of Alzheimer's disease are screened and verified by a variety of means. They are committed to repeated validation to ensure the accuracy of the results and identify credible biomarkers from the candidate biomarkers. Methods: the first part of the study was based on AD and healthy controls, using enzyme linked immunosorbent assay. The enzyme linked immunosorbent assay (ELISA) verifies the AD biomarkers reported in the previous literature, including Apolipoprotein A1, Haptoglobin, Alpha-1-antitrypsin, Clusterin, C-reative statements, and second parts. For the study, the biomarkers of Alzheimer's disease were studied by means of isobaric tags for relative and absolute quantitation, iTRAQ and protein chip technology, combined with bioinformatics analysis, and some differential proteins were tested by ELISA and immunological trace technology (Western blot, WB). Evidence, including: Afamin, Platelet basic protein, Fibulin-1, Plasma protease C1inhibitor, Thrombospondin-1, Inter-alpha-trypsin inhibitor heavy, and healthy control as the research object, using the technique to study the two groups of blood differential proteins. Results: the first part Some of the biomarkers in the literature were verified. The expression level of 6 proteins was statistically significant, in which Haptoglobin was significantly up-regulated, Apolipoprotein A1, Alpha-1-antitrypsin, Matrix metalloproteinase 9, Calgranulin A and Calgranulin B decreased significantly. The 6 proteins were Apolipoprotein A1. The trend of B is consistent with the literature report. In the second part, 7 proteins were screened by iTRAQ and protein chip technology, of which Platelet basic protein, Fibulin-1 and Fibrinogen gamma chain were significantly up-regulated, Plasma protease C1. The trend of 1 inhibitor is consistent with the trend of literature report and iTRAQ result, the change trend of Fibulin-1 and Thrombospondin-1 is the same as that of iTRAQ. The change trend of Ceruloplasmin and Fibrinogen gamma chain is consistent with the literature report, the result of protein chip and WB results. The third part is the experiment object of the control group and the MCI group, and the iTRAQ technology is used. Conclusion: the results of this study suggest that Alzheimer's disease may be associated with the changes in the level of expression of Apolipoprotein A1, Calgranulin A, Calgranulin B, Plasma protease C1 inhibitor, Thrombospondin-1, Ceruloplasmin, and such proteins may be potential organisms for Alzheimer's disease. Markers. Whether these proteins can be used as biomarkers for Alzheimer's disease need further study.
【学位授予单位】：深圳大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R749.16

【相似文献】