CUMS抑郁模型小鼠肝脏的iTRAQ定量蛋白质组学研究

发布时间：2018-05-06 03:10

本文选题：抑郁症 + 慢性不可预知性温和刺激(CUMS)　；参考：《重庆医科大学》2017年硕士论文

【摘要】：研究背景抑郁症是一种高度流行且致残的精神疾病,对人们的生活质量和机体功能有重大损害。然而,我们对抑郁表型及其潜在的病理生理学机制之间的关系仍然知之甚少。现阶段对抑郁症的研究主要集中在脑区,脑与机体的联合探索可能会为抑郁症病因的全面理解提供新的线索。作为人体最大的内脏器官,肝脏在机体生物转化,合成,代谢等一系列生理和生化反应中发挥着重要的作用。但目前仍然没有用蛋白质组学方法对抑郁模型小鼠肝脏中蛋白表达进行评估的相关研究。目的本研究旨在用iTRAQ标记和LC-MS/MS联合的蛋白质组学技术鉴定CUMS抑郁模型小鼠肝脏的蛋白表达谱,用特定的筛选条件寻找相关的差异蛋白,初步分析这些差异蛋白及其所涉及通路的病理生理机制,探讨抑郁状态下肝脏结构功能改变与蛋白表达谱变化的关系,为诊治抑郁症提供新的思路和方向。方法经过环境适应和一周的糖水训练之后,我们随机地把40只正常的C57BL/6J雄性小鼠分为抑郁模型组和正常对照组(20:20)。20只模型组小鼠进行4周的慢性不可预知性温和刺激之后,将两组进行行为学评估,若模型成立,从每组中随机选取小鼠肝脏组织(10:10)进行itraq蛋白组学鉴定,对差异蛋白进行生物信息学分析,另每组选取6只小鼠肝脏组织对差异蛋白和通路进行免疫印迹法(Western blot,WB)验证。结果1.模型组CUMS之后对两组的体重,糖水偏好,强迫游泳,旷场试验等行为学数据进行统计学分析,有显著差异,证明造模成功。2.Itraq串联质谱鉴定出4000多种蛋白质,用特殊肽段大于等于2,1.2以上差异倍数,P值小于0.05三个条件,筛选出66种最显著差异的蛋白质,用于进一步的生物信息学分析。3.通过信号网络分析(IPA),我们发现这66种差异蛋白与炎症反应,免疫调节,脂质代谢和NFκB信号通路等病理生理过程相关。4.此外,通过免疫印迹法(Western blot,WB)验证了与这些通路密切相关的四种蛋白质,血红蛋白(HPX),触珠蛋白(HP),细胞色素P4502A4(CYP2A4)和胆汁盐磺基转移酶1(SULT2A1),结果与itraq一致。结论我们的研究首次报道了CUMS抑郁模型小鼠中肝脏组织的蛋白表达谱,为以后研究抑郁症多层面的机制提供了如肝-脑轴等新的见解。
[Abstract]:Background Depression is a highly prevalent and disabling mental disease, which has great damage to people's quality of life and body function. However, little is known about the relationship between depression phenotypes and their underlying pathophysiological mechanisms. At present, the research on depression is mainly focused on the brain area. The combined exploration of brain and body may provide new clues for the comprehensive understanding of the cause of depression. As the largest visceral organ, liver plays an important role in biological transformation, synthesis, metabolism and a series of physiological and biochemical reactions. However, no proteomics method has been used to evaluate the expression of protein in the liver of depressive mice. Objective to identify the liver protein expression profiles of CUMS depressive mice by iTRAQ labeling and LC-MS/MS proteomics, and to search for differential proteins by specific screening conditions. The pathophysiological mechanisms of these differentially expressed proteins and their related pathways were preliminarily analyzed, and the relationship between the changes of liver structure and function and the changes of protein expression profile in depressive state was discussed, which provided a new way of thinking and direction for the diagnosis and treatment of depression. Methods after environmental adaptation and a week of sugar water training, we randomly divided 40 normal C57BL/6J male mice into depression model group and normal control group 20: 20 mice. After 4 weeks of chronic unpredictable mild stimulation, 40 normal male C57BL/6J mice were randomly divided into depression model group and normal control group. The behavior of the two groups was evaluated. If the model was established, the liver tissues of each group were randomly selected for itraq proteomics identification, and the differential proteins were analyzed by bioinformatics. In addition, the liver tissues of 6 mice in each group were examined by Western blotWB for differential protein and pathway. Result 1. After CUMS, the behavioral data such as body weight, sugar water preference, forced swimming, open field test and so on in the model group were statistically analyzed. The results showed that the model was successful. 2. Itraq tandem mass spectrometry identified more than 4000 proteins. Using the three conditions that the special peptide segment is larger than or equal to 2o 1.2 and the difference multiple P value is less than 0.05, 66 proteins with the most significant difference are selected for further bioinformatics analysis. By signal network analysis, we found that these 66 differential proteins were related to inflammatory response, immune regulation, lipid metabolism and NF 魏 B signaling pathway. In addition, four proteins closely related to these pathways, HPX, HPX, cytochrome P4502A4 (CYP2A4) and bile salt sulfotransferase 1 (SULT2A1), were identified by Western blotDNA (Western blot). The results were consistent with those of itraq. Conclusion our study reported for the first time the expression profile of liver protein in CUMS depression model mice, which provided some new insights into the mechanism of depression in the future, such as liver and brain axis.
【学位授予单位】：重庆医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R749.4;R-332

【参考文献】