Proteomic analysis of the hippocampus in Alzheimer's disease model mice by using two-dimensional fluorescence difference in gel electrophoresis
Proteomic analysis of the hippocampus in Alzheimer's disease model mice by using two-dimensional fluorescence difference in gel electrophoresis
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DOI：
10.1016/j.neulet.2012.11.010
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发表时间：
2013-02-08
影响因子：
2.5
通讯作者：
Matsuyama, Shogo
中科院分区：
文献类型：
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作者：
Takano, Masaoki;Yamashita, Takuya;Matsuyama, Shogo
我们先前在阿尔茨海默病（AD）患者中鉴定了淀粉样前体蛋白（APP）中的E693 Δ突变，然后产生表达该突变的APP转基因小鼠。由于这些小鼠从8月龄开始就具有丰富的A β寡聚体，但即使在24月龄也没有淀粉样蛋白斑块，因此它们是研究淀粉样蛋白β（A β）寡聚体的病理作用的良好模型。凝胶电泳中的二维荧光差异（2D-DIGE）技术，使用混合样品内标，现在被认为是一种准确的方法来确定和定量蛋白质。在这项研究中，我们研究了蛋白质的水平改变，在12个月大的APP（E693德尔塔）转基因小鼠的海马使用二维DIGE和液相色谱-串联质谱（LC-MS/MS）。在APP（E693 Delta）转基因小鼠的海马中有14种蛋白质发生了显著变化。肌动蛋白细胞质1（β-肌动蛋白）、热休克同源物71 kDa、γ-烯醇化酶、ATP合酶β亚基、微管蛋白β-2A链、网格蛋白轻链B（网格蛋白）和发动蛋白-1增加。热休克相关的70 kDa蛋白2、神经丝轻多肽（NFL）、应激诱导的磷蛋白2、60 kDa热休克蛋白（HSP 60）、α-内连接蛋白、蛋白激酶C和神经元蛋白1（Pacsin 1）中的酪蛋白激酶底物、α-烯醇化酶和β-肌动蛋白减少。Western blotting也验证了APP（E693 Delta）转基因小鼠中HSP 60、NFL、网格蛋白和Pacsin 1水平的变化。所鉴定的蛋白质可分为细胞骨架蛋白、分子伴侣蛋白、神经传递蛋白、能量供应蛋白和信号转导蛋白。因此，通过2D-DIGE和LC-MS/MS的蛋白质组学提供了AD脑早期阶段蛋白质水平的知识。(C)2012爱思唯尔爱尔兰有限公司保留所有权利。
We previously identified the E693 Delta, mutation in amyloid precursor protein (APP) in patients with Alzheimer's disease (AD) and then generated APP-transgenic mice expressing this mutation. As these mice possessed abundant A beta oligomers from 8 months of age but no amyloid plaques even at 24 months of age, they are a good model to study pathological effects of amyloid beta (A beta) oligomers. The two-dimensional fluorescence difference in gel electrophoresis (2D-DIGE) technology, using a mixed-sample internal standard, is now recognized as an accurate method to determine and quantify proteins. In this study, we examined the proteins for which levels were altered in the hippocampus of 12-month-old APP(E693 Delta)-transgenic mice using 2D-DIGE and liquid chromatography-tandem mass spectrometry (LC-MS/MS). Fourteen proteins were significantly changed in the hippocampus of APP(E693 Delta)-transgenic mice. Actin cytoplasmic 1 (beta-actin), heat shock cognate 71 kDa, gamma-enolase, ATP synthase subunit beta, tubulin beta-2A chain, clathrin light chain B (clathrin) and dynamin-1 were increased. Heat shock-related 70 kDa protein 2, neurofilament light polypeptide (NFL), stress-induced-phosphoprotein 2, 60 kDa heat shock protein (HSP60), alpha-internexin, protein kinase C and casein kinase substrate in neurons protein 1 (Pacsin 1), alpha-enolase and beta-actin were decreased. Western blotting also validated the changed levels of HSP60, NFL, clathrin and Pacsin 1 in APP(E693 Delta)-transgenic mice. The identified proteins could be classified as cytoskeleton, chaperons, neurotransmission, energy supply and signal transduction. Thus, proteomics by 2D-DIGE and LC-MS/MS has provided knowledge of the levels of proteins in the early stages of AD brain. (C) 2012 Elsevier Ireland Ltd. All rights reserved.