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研究生:葉心卉
研究生(外文):Sin-Huei Yeh
論文名稱:葡萄糖胺調控腦神經細胞中神經可塑性相關基因與蛋白之表現
論文名稱(外文):Glucosamine regulation of neuroplasticity-related gene and protein expression in brain neuronal cells
指導教授:吳鈺琳
指導教授(外文):Wu, Yuh-Lin
口試委員:楊定一鄭瓊娟李怡萱
口試委員(外文):Yang, Ding-YiJeng, Chung-JiuanLi, Yi-Hsuan
口試日期:2022-01-19
學位類別:碩士
校院名稱:國立陽明交通大學
系所名稱:生理學研究所
學門:醫藥衛生學門
學類:醫學學類
論文種類:學術論文
論文出版年:2022
畢業學年度:110
語文別:中文
論文頁數:71
中文關鍵詞:阿茲海默症β-澱粉樣蛋白葡萄糖胺神經可塑性學習記憶
外文關鍵詞:Alzheimer’s diseaseGlucosamineNeuroplasticityLearning and memory
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中文摘要 i
Abstract iii
目錄 v
圖目錄 vii
壹、前言 (Introduction) 1
一、記憶的形成 1
二、神經可塑性因子調節神經系統和鞏固記憶 1
三、生長相關蛋白 (growth-associated protein 43, GAP-43)在神經細胞調節中的角色 3
四、腦源性神經滋養因子 (Brain-derived neurotrophic factor, BDNF)以及成纖維細胞生長因子21 (Fibroblast growth factor 21, FGF21)在神經系統中的重要性 4
五、腦部神經功能受損與退化性疾病 5
六、Glucosamine的生理角色 7
七、研究動機 8
貳、實驗材料與方法 (Materials and methods) 10
一、實驗藥品及試劑 (Chemicals and reagents) 10
二、動物實驗 12
1.實驗動物 (Animal model) 12
2.動物行為實驗 (Animal behavior experimentation) 13
三、細胞培養 (Cell culture) 13
1.HT22 cell line: 13
2.初代皮質層神經元 (Primary cortical neuron) 14
四、Aβ25-35製備 (Aβ25-35 preparation) 14
五、細胞存活率測試 (Cell viability assay) 15
六、酵素結合免疫吸附分析法(Enzyme-linked immunosorbent assay, ELISA) 15
七、反轉錄聚合酶鏈鎖反應 (Reverse transcription-polymerase chain reaction, RT-PCR) 16
1.去氧核醣核酸 (RNA)萃取: 16
2.反轉錄作用製備 (cDNA synthesis) 16
3.聚合酶鏈鎖反應 (Polymerase chain reaction) 17
八、西方墨點法 (Western blotting) 19
1.蛋白質萃取: 19
2.蛋白質變性膠體電泳 (sodium dodecyl sulphate–polyacrylamide gel electrophoresis, SDS-PAGE): 19
3.漬膜轉移 (Transfer): 20
4.抗體作用(antibody reaction): 21
5.冷光螢光影像擷取系統 (GE Amersham Imager 600): 22
九、免疫螢光染色 (Immunofluorescence, IF): 22
1.細胞準備和固定: 22
2.抗體作用 (Antibody reaction): 23
3.正立式螢光顯微鏡 (Olympus BX61): 24
十、統計分析 (Statistical analysis): 24
參、結果 (Results) 25
一、葡萄糖胺促進小鼠的學習與記憶功能 25
二、 葡萄糖胺促進小鼠海馬迴的SYP、PSD-95、Densin-180的mRNA或蛋白表現量增加 25
三、葡萄糖胺促進海馬迴的GAP-43、BDNF、FGF21的mRNA和蛋白的表現 25
四、葡萄糖胺促進小鼠皮質層的SYP、PSD-95、Densin-180的mRNA或蛋白表現 26
五、葡萄糖胺促進小鼠皮質層的GAP-43、BDNF、FGF21、GDNF、IL-10的mRNA或蛋白表現 27
六、葡萄糖胺在海馬迴神經細胞株HT22中並不會影響細胞存活率並且促進SYP、PSD-95、Densin-180的mRNA或蛋白表現 28
七、葡萄糖胺在海馬迴神經細胞株HT22中促進GAP-43和FGF21的蛋白表現量以及GDNF的mRNA表現 28
八、葡萄糖胺在大鼠初代皮質神經細胞中並不會影響細胞存活率並且促進SYP、Densin-180的mRNA或蛋白表現 29
九、葡萄糖胺在初代皮質神經細胞中促進GAP-43和BDNF的mRNA和蛋白表現 30
十、葡萄糖胺在海馬迴細胞株HT22中增強SYP之表現 30
十一、葡萄糖胺在大鼠初代皮質神經細胞中增強SYP、GAP-43在突觸位置的表現量 31
十二、Aβ (amyloid beta)導致HT22以及初代皮質神經細胞的細胞存活率下降 32
十三、Aβ25-35在海馬迴神經細胞HT22中對於SYP、PSD-95、Densin-180以及GAP-43、BDNF、FGF21的mRNA和蛋白表現的影響 32
肆、討論 (Discussion) 33
伍、參考文獻 (References) 37
陸、圖與圖誌 (Figures and Legends) 43

圖目錄
圖一、葡萄糖胺促進小鼠的學習記憶功能。 43
圖二、葡萄糖胺促進小鼠海馬迴中SYP、PSD-95、Densin-180之mRNA或蛋白表現。 45
圖三、葡萄糖胺促進小鼠海馬迴中GAP-43、FGF21、BDNF的mRNA和蛋白表現。 47
圖四、葡萄糖胺促進小鼠皮質層中SYP、PSD-95、Densin-180之mRNA和蛋白表現。 49
圖五、葡萄糖胺促進小鼠皮質層中GAP-43、BDNF、FGF21、GDNF、IL-10的mRNA或蛋白表現。 51
圖六、葡萄糖胺對於海馬迴神經細胞HT22的細胞存活率之影響以及促進SYP、PSD-95、Densin-180之mRNA或蛋白表現量。 53
圖七、葡萄糖胺對於海馬迴神經細胞HT22中GAP-43、BDNF、FGF21、GDNF之mRNA或蛋白表現以及FGF21分泌的含量之影響。 55
圖八、葡萄糖胺對初代皮質神經細胞的細胞存活率以及SYP、PSD-95、Densin-180之mRNA或蛋白表現之影響。 58
圖九、葡萄糖胺對於初代皮質神經細胞中GAP-43、BDNF、FGF21的mRNA和蛋白表現之影響。 60
圖十、葡萄糖胺在海馬迴神經細胞HT22中SYP、PSD95、Densin-180、GAP-43的分佈位置以及表現的影響。 62
圖十一、葡萄糖胺在初代皮質神經細胞中SYP、PSD95、GAP-43的分佈位置以及表現的影響。 65
圖十二、Aβ造成海馬迴神經細胞HT22和初代皮質神經細胞的細胞存活率下降。 66
圖十三、Aβ和葡萄糖胺對於海馬迴神經細胞HT22中SYP、PSD-95、Densin-180、GAP-43、BDNF、FGF21的mRNA和蛋白表現之影響以及FGF21分泌的含量之影響 69
附圖一、葡萄糖胺對小鼠紋狀體中SYP、PSD-95、Densin-180、GAP-43、BDNF、FGF21、GDNF和IL-10的mRNA表現。 71
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