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研究生:王亮博
研究生(外文):Wang, Liang-Bo
論文名稱:探討小分子C-2對Protein-X促進amyloid beta寡聚合及細胞毒性之研究
論文名稱(外文):Investigation of a small molecule, C-2, for its anti- Protein-X inducing amyloid beta oligomerization and cytotoxicity
指導教授:林榮耀林榮耀引用關係賴韻如賴韻如引用關係
指導教授(外文):Lin, Jung-YawLai, Yun-Ju
學位類別:碩士
校院名稱:國立臺灣師範大學
系所名稱:生命科學系
學門:生命科學學門
學類:生物學類
論文出版年:2020
畢業學年度:109
語文別:英文
論文頁數:39
中文關鍵詞:阿茲海默氏症X蛋白類澱粉蛋白-β肽
外文關鍵詞:Alzheimer’s disease (AD)Protein-Xβ-amyloid (Aβ)
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阿茲海默氏症(Alzheimer’s disease, AD)已被視為常見的神經退化性疾病,主要病因為海馬迴的神經元受損,同時也引起認知功能障礙的出現。AD特徵包括由細胞內蛋白tau過度磷酸化所造成的神經纖維糾結(neurofibrillary tangles, NFTs)及細胞外類澱粉蛋白-β肽(β-amyloid peptide, Aβ)聚集形成的類澱粉斑塊(plaques)。目前,我們實驗室已發現,與年幼的小鼠相比,年老的AD模式鼠具有較高表現量的X蛋白。為了更進一步了解X蛋白在AD中的作用機制,我們藉由轉殖入X蛋白的SH-SY5Y細胞株作為本實驗使用的模式細胞。在細胞存活率方面以及西方墨點法的實驗中,從寡聚合物Aβ42中處理的細胞組別可以得知,經X蛋白的給予後會產生更多Aβ的聚集,使得細胞存活率降低,而處理小分子藥物的C-2能有效降低X蛋白所給予出的Aβ,進而達到拯救細胞的效果。因此,我們認為C-2可能透過抑制X蛋白而有效治療AD的小分子藥物。
Alzheimer’s disease (AD) has been considered as a common neurodegenerative disorder that causes hippocampal neurons damages and cognitive dysfunctions. AD features include neurofibrillary tangles (NFTs) caused by hyper-phosphorylation of intracellular tau protein, and extracellular β-amyloid (Aβ) plaques by Aβ aggregation. Our laboratory recently found that Protein-X level and the oligomerization of Aβ were concomitantly increased in aged triple transgenic AD mice (3xTg-AD) [PS1M146V, APPK670M/N671L, and tauP301L]. To further understand the mechanism of Protein-X in AD pathway, we established Protein-X stable-transfected SH-SY5Y cell line as the cell model in this experiment. In cell-based system and western blot analysis, Protein-X enhanced the formation of Aβ aggregation and the cytotoxicity is more than that in the present of Protein-X, implying that this cell-based system could serve as a platform for drug screening. Furthermore, we found that the small molecule, C-2, had the ability decrease the Aβ42 cytotoxicity induced by Protein-X. C-2 could be anticipated to develop a promising therapeutic agent for AD.
Introduction 1
1. Alzheimer’s disease 1
2. β-amyloid (Aβ) formation 2
3. The relationship between Aβ and neuronal death 2
4. AD animal model, triple transgenic mice, (3xTg-AD) 3
5. Research aims 3


Materials and Methods 4
1. Materials 4
2. Cell culture 5
3. Cell viability assay 5
3.1 Preparation of Aβ42 for Cell viability assay 6
4. Western blotting 6
4.1 Preparation of cell lysates 6
4.2 Quantification of protein concentration 6
5. Detemination of Aβ42 oligomerization in the cell-free system by Western blot analysis 7
5.1 Preparation of the cell-free protein sample 7
5.2 Aβ42 oligomerization in the cell-free system by Western blot analysis 7
6. Animals 8
6.1 Morris water maze test 8
6.2 Y maze task 8
6.3 Novel object recognition experiment 9
6.4 Statistical analysis 9


Results 10
1. Protein-X enhanced the formation of Aβ42 aggregation with senescence in the brain of 3xTg-AD mice 10
2. Effects of the small molecule, C-2 on Aβ42 cytotoxicity that enhanced by Protein-X 10
3. C-2 reduced Aβ42 oligomerization induced by Protein-X in the cell-free system 11
4. Treatment with the small molecule, C-2, downregulated cell apoptosis markers induced by Aβ42 12
5. Treatment with the small molecule, C-2, downregulated tau hyper-phosphorylation markers induced by Aβ42 12
6. C-2 upregulated tau hyper-phosphorylation regulator, GSK-3β 13
7. C-2 improves the learning and memory deficits of 3xTg-AD mice 13
8. C-2 treatment ameliorates the behavior on the novel object recognition task in 3xTg-AD mice 14
9. C-2 treatment significantly reduced memory behaviors in the Y maze task 14


Discussion 15
References 18
Figures 22
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