ApoE4对突触线粒体损伤和氧化应激水平的影响与Alzheimer病

发布时间：2018-03-16 05:12

本文选题：载脂蛋白E氧化应激线粒体突触体阿尔兹海默病　切入点：　出处：《中国科学技术大学》2012年博士论文　论文类型：学位论文

【摘要】：载脂蛋白E4(ApoE4),在人群中有14%的等位基因频率。与ApoE3基因相比,它作为一个风险因子,被认为在散发性阿尔兹海默病中起着重要的作用。近年来研究显示,氧化应激和突触功能结构上的改变在阿尔兹海默病病理过程中发挥了重要的作用,这三者之间密切相关。我们的研究就是基于这个基础,希望可以发现在阿尔兹海默病的病理过程中,ApoE的基因多态性是如何通过氧化应激和突触而发挥作用的我们选择了13个月大的ApoE转基因鼠作为阿尔兹海默病模型鼠。我们对其进行了行为学测试,结果显示ApoE4转基因鼠在30分钟短间隔的新奇物体识别任务中,表现出了较差的学习认知能力。而在Y迷宫任务中,无论是短间隔任务还是长间隔任务,ApoE4型转基因鼠相比于ApoE3型并没有表现出认知功能的损伤。我们利用percoll梯度离心的方法成功分离了突触体,并鉴定了它的纯度。将分离的突触体进行了定量的蛋白质组学分析。我们发现,在13个月龄的ApoE4型转基因鼠中,涉及糖类代谢,氧化应激和电子传递呼吸链的蛋白表达水平明显下降。丙二醛(MDA)是活性氧自由基(ROS)攻击多聚不饱和脂肪酸的产物。它被认为和谷胱甘肽系统一样,是衡量体内氧化应激水平的一个可靠标志物。我们发现在ApoE4型转基因鼠的皮层组织中,丙二醛和氧化型谷胱甘肽的水平显著提高。进一步分析发现作为衡量谷胱甘肽系统稳定性的还原型谷胱甘肽/氧化型谷胱甘肽比值在ApoE4型小鼠中显著下降。这表明13个月的ApoE4转基因鼠处在一个高氧化应激水平中。为了验证我们蛋白质组学的结果。我们检测了超氧化物歧化酶2(SOD2)和细胞色素C氧化酶亚基4(COX IV)的蛋白水平表达。这是2个在氧化应激和线粒体电子传递呼吸链上发挥着重要作用的蛋白。与之前蛋白质组学的分析结果相一致。我们发现在13月龄的ApoE4型转基因小鼠皮层中,他们的表达水平明显降低。与蛋白质组学和Western结果所显示的相关蛋白水平差异不同,我们并没有发现涉及突触可塑性,氧化应激以及线粒体功能相关基因的mRNA水平有表达差异。这可能提示我们ApoE对于氧化应激和线粒体的调控并不是通过转录水平实现的。综上所述,我们的研究提示了在ApoE转基因鼠中存在着ApoE4型依赖的突触线粒体损伤和氧化应激水平提高,这可能是阿尔兹海默病病理过程中的重要诱因。
[Abstract]:Apolipoprotein E4 (ApoE4), there are 14% alleles in the population frequency. Compared with the ApoE3 gene, as a risk factor, are believed to play an important role in sporadic Alzheimer's disease. Recent studies have shown that oxidative stress and synaptic structural changes play an important role in the pathological process of Alzheimer's disease, is closely related to the three. Our research is based on this foundation, hope can be found in the pathological process of Alzheimer's disease, ApoE gene polymorphism is how to play a role through oxidative stress and synapse
We chose the 13 month old ApoE transgenic mice as a model of Alzheimer's disease rats. We carried on the behavioral test, the results showed that ApoE4 transgenic mice in the novel object recognition task 30 minutes short interval, showed poor cognitive ability. Learning in the Y maze task, whether it is a short interval the task or long interval task, compared to ApoE4 in ApoE3 transgenic mice did not exhibit cognitive dysfunction.
We use the method of Percoll gradient centrifugation were successfully separated and identified synaptosomes, its purity. The isolated synaptosomes were quantitative proteomics analysis. We found that in ApoE4 transgenic mice 13 months of age, involved in carbohydrate metabolism, oxidative stress and the expression level of electron transport in the respiratory chain protein decreased significantly.
Malondialdehyde (MDA) is the active oxygen free radicals (ROS) attack polyunsaturated fatty acid product. It is considered as the glutathione system, is a reliable indicator of the level of oxidative stress in vivo. We found that in ApoE4 transgenic mice cortex, malondialdehyde and oxidized glutathione levels increased significantly further analysis found that the stability of the system as a measure of glutathione GSH / GSSG ratio decreased significantly in ApoE4 mice. This indicates that 13 months of ApoE4 transgenic mice at a high level of oxidative stress.
In order to verify our proteomics results. We measured superoxide dismutase 2 (SOD2) and cytochrome C oxidase subunit 4 (COX IV) protein expression level. This is the 2 transfer of breathing in the oxidative stress and mitochondrial electron chain plays an important role in the analysis of protein and protein before. Proteomics results. We found that in ApoE4 transgenic mice cortex 13 month old, their expression levels were significantly reduced.
Correlation between protein level display and proteomics and Western results, we have not found involved in synaptic plasticity, oxidative stress and mitochondrial function related gene mRNA expression difference. This may suggest that ApoE on oxidative stress and mitochondrial regulation is not by the transcription level.
In conclusion, our research suggests that there are ApoE4 dependent mitochondrial damage and oxidative stress levels in ApoE transgenic mice, which may be an important inducement of Alzheimer's disease.

【学位授予单位】：中国科学技术大学
【学位级别】：博士
【学位授予年份】：2012
【分类号】：R749.16

【共引文献】