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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 蔡蔭和(Inn-Ho Tsai) | |
| dc.contributor.author | Yung-Chih Cheng | en |
| dc.contributor.author | 鄭永志 | zh_TW |
| dc.date.accessioned | 2021-06-13T02:18:38Z | - |
| dc.date.available | 2021-08-01 | |
| dc.date.copyright | 2011-08-05 | |
| dc.date.issued | 2011 | |
| dc.date.submitted | 2011-08-01 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/30863 | - |
| dc.description.abstract | 阿茲罕莫症、帕金森氏症與亨丁頓舞蹈症等神經退化疾病經常引發中樞神經系統內神經細胞死亡，或造成神經細胞喪失功能 。截至目前，造成神經細胞死亡的原因仍不明朗化 。對於如何避免神經細胞走向死亡的路徑，向來是研究神經相關疾病的重要議題 。在這裡我們強調底板蛋白F-spondin對於神經細胞的保護功能，以及它本身避免神經細胞死亡的潛在能力。
我們發現3’末端具有多個鳥嘌呤的寡去氧核苷酸擁有引發胚胎神經細胞死亡和抑制神經突延伸的能力，且轉錄因子Ybox一號固定蛋白也被鑑定出在胚胎神經細胞的細胞質中，會和這種寡去氧核苷酸進行連結，而連結後的複合體會促進Ybox一號固定蛋白大量進入胚胎神經細胞的細胞核 。此外，我們也發現在細胞核內的Ybox一號固定蛋白會接合於底板蛋白的起動器區域，並且抑制底板蛋白的表現。同時證實了3’末端具有多個鳥嘌呤的寡去氧核苷酸造成神經細胞死亡的現象，主要是由於大量的Ybox一號固定蛋白進入細胞核，並且抑制底板蛋白F-spondin的表現所造成的 。當我們去除胚胎神經細胞內Ybox一號固定蛋白的細胞核移位序列，或在神經細胞內大量表現底板蛋白，可以有效減少此種寡核苷酸所造成的神經細胞死亡現象，並且回復神經突應有的長度。 另外，亦發現在老鼠的神經瘤細胞株會分泌一定量的第六號介白素 。利用小片段干擾RNA的技術，將底板蛋白的表現量降低，會造成老鼠神經瘤本身的第六號介白素降低，並且發現對於血清營養不足或類澱粉蛋白胜肽片段所造成的細胞毒殺作用比較沒有抵抗能力 。而當加入底板蛋白的重組蛋白、第六號介白素重組蛋白，或大量在胚胎神經細胞內表現底板蛋白，皆可使因為小片段干擾RNA所減少的底板蛋白重新回復，並且也使得因為血清營養不足或類澱粉蛋白胜肽片段所造成的細胞死亡數量大量減少 。我們也知道第六號介白素的降低，是由於胚胎神經細胞當中核因子 NF-κB的活性降低，同時也和 p38有絲分裂活化蛋白激酶的抑制現象呈正相關。大量表現 p38有絲分裂活化蛋白激酶的活化蛋白，可增加第六號介白素的表現量，同時也回復因為底板蛋白的小片段干擾RNA所降低的p38有絲分裂活化蛋白激酶活性，並且減少因為類澱粉蛋白胜肽片段所造成的神經細胞死亡現象 。總而言之，這些結果發現胚胎神經細胞與老鼠神經瘤內的底板蛋白，是一個關乎細胞存活的重要關鍵。 | zh_TW |
| dc.description.abstract | Neurodegenerative diseases, including Alzheimer’s (AD), Parkinson’s (PD) and Huntington’s disease, always trigger a massive neuronal cell death or lead to neuronal dysfunction in the central nervous system. However, the exact causes of this neuronal loss are still unclear. How to protect neurons from cell death is an important issue in the fight against these neurological diseases. Herein, we highlight the protective role of the protein F-spondin protein and its potential therapeutic implications in preventing neuronal cell death.
We found that oligodeoxynucleotides containing poly-guanines at the 3’ terminus (3’G-ODN) have the ability to induce cell death and to inhibit neurite extension in embryonic neuron cells. The transcription factor YB-1 in the cytoplasm was identified to associate with 3’G-ODN and this associated complex enhanced the translocation of YB-1 into the nucleus. Once in the nucleus YB-1 then binds to the promoter of F-spondin and represses the expression of F-spondin. Deletion of the nuclear localizing sequence (NLS) of YB-1 or overexpression of F-spondin in embryonic neuron cells rescued the cell death and neurite retraction induced by 3’G-ODN in embryonic neuron cells. In addition, we found that murine neuroblastoma cells expressed a significant level of interleukin-6 (IL-6). Knock-down of F-spondin mRNA in murine neuroblastoma cells using small interfering RNAs led to decreased IL-6 levels along with lower resistance to serum starvation and cytotoxic amyloid β1-42 (Aβ1-42) peptide. Adding exogenous F-spondin or IL-6 recombinant protein, even in over-expressing F-spondin in murine neuroblastoma cells, restored the decline of F-spondin or IL-6 expression induced by F-spondin knockdown and reversed the cell death induced by Aβ1-42 peptide or serum starvation. The decrease in the level of IL-6 level was positively correlated with a decrease in the level of NF-κB and inhibition of p38 mitogen-activated protein kinase (MAPK). Over-expressing MEKK, a kinase activator of the p38 MAPK pathway, increased IL-6 production, restored the decrease of p38 induced by F-spondin knock-down, and rescued the cells from death caused by Aβ1-42 peptide. Taken together, these results suggest that F-spondin is an important protein in responsible for cell survival in embryonic neuron cells and murine neuroblastoam cells. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-13T02:18:38Z (GMT). No. of bitstreams: 1 ntu-100-D94b46006-1.pdf: 27923183 bytes, checksum: 86d7f3d14b90e12fc6caab4f32abd8ea (MD5) Previous issue date: 2011 | en |
| dc.description.tableofcontents | Table of Contents
口試委員會審訂書 i 謝 辭 ii 中文摘要 iv Abstract vi List of Appendices xv Abbreviations xvi Introduction 1 1. Central nervous system (CNS) 1 2. Neurodegenerative diseases 2 2.1. Infections and neurodegeneration 4 2.2. Alzheimer’s disease (AD) 5 2.3. Neuronal cell death 6 3. Toll-like receptor (TLR) family 7 3.1. TLR signaling 9 3.2 TLR expression in cells of the brain 10 4. CpG-Oligodeoxynucleotides (CpG-ODNs) 11 5. G-quadruplex structure 13 6. F-spondin 15 7. Y-Box binding protein – 1 (YB-1) 18 8. Interleukin-6 (IL-6) 18 Hypothesis 20 Aims and Study Approaches 21 Materials and Methods 22 1. Animals 22 2. Reagents 22 3. Cell culture 23 4. Cytokine- and chemokine- specific ELISA 24 5. Detection of TLR expression in embryonic neuron cells 24 6. Isolation and culture of primary embryonic neuron cells 25 7. Transfection of embryonic neuron cells 26 8. Neurite outgrowth analysis 27 9. Cell death and viability assay 27 10. Caspase enzymatic activity assay 28 11. Analysis of ODN uptake 28 12. Pull-down assay 29 13. RNA isolation and real-time quantitative PCR 29 14. Western immunoblotting 30 15. Plasmid construction and transient transfection 31 17. Stealth siRNA knock-down 33 18. Generation of F-spondin-silenced stable cell lines 33 19. Nitric oxide assay 34 20. Luciferase reporter assay 34 21. Statistical analysis 34 Results 36 1. The expression of TLRs on embryonic neuron cells 36 2. Effects of TLR ligands on embryonic neuron cells 36 3. The effect induced by CpG-A ODN is not mediated by TLR 9 37 4. 3’G-ODN induces neuron cell aggregation and neurite retraction 37 5. 3’G-ODN induces neuron cell death through caspase 3/7 pathways 38 6. 3’G-ODNs enter the cytoplasm and associate with YBOX1 protein in embryonic neuron cells 39 7. Stimulation of 3’G-ODNs enhances nuclear translocation of YB-1 41 8. F-spondin expression is repressed by 3’G-ODNs in embryonic neuron cells 42 9. YB-1 translocates to the nucleus and represses the transcription of F-spondin 43 10. A YB-1 mutant lacking the NLS results in insensitivity of embryonic neuron cells to cytotoxicity induced by 3’G-ODN 43 11. Ectopic expression of F-spondin protein renders embryonic neuron cells more resistant to cytotoxic 3’G-ODN 45 12. Expression of F-spondin differs between embryonic neuron and neuroblastoma cells 46 13. Association of F-spondin with IL-6 expression in murine neuroblastoma cells 46 14. Prevention of cytotoxic stimuli-mediated cell death by F-spondin and IL-6 47 15. F-spondin gene knockdown decreases NF-κB activity 49 16. Maintenance of IL-6 expression by F-spondin is associated with p38 MAPK 49 17. Ectopic expression of constitutively active MEK kinase activates p38 MAPK and renders Neuro-2aSpondinRNAi cells more resistant to cytotoxic Aβ1-42 peptide 50 Discussion 52 | |
| dc.language.iso | en | |
| dc.subject | 神經疾病 | zh_TW |
| dc.subject | 底板反應蛋白 | zh_TW |
| dc.subject | G-四聯體 | zh_TW |
| dc.subject | 寡去氧核苷 | zh_TW |
| dc.subject | 酸 | zh_TW |
| dc.subject | 胚胎神經細胞 | zh_TW |
| dc.subject | YBox 一號 結合蛋白 | zh_TW |
| dc.subject | 第六號介白素 | zh_TW |
| dc.subject | p38有絲分裂活化蛋白激酶 | zh_TW |
| dc.subject | 神經細胞瘤 | zh_TW |
| dc.subject | 神經細胞存活 | zh_TW |
| dc.subject | 神經退化疾病 | zh_TW |
| dc.subject | neuronal survival | en |
| dc.subject | p38 MAPK | en |
| dc.subject | embryonic neuron cell | en |
| dc.subject | neuroblastoma | en |
| dc.subject | neurodegenerative disease | en |
| dc.subject | neurological disorders | en |
| dc.subject | F-spondin | en |
| dc.subject | G-quadruplex | en |
| dc.subject | oligodeoxynucleotide | en |
| dc.subject | Y box binding protein | en |
| dc.subject | interleukin-6 | en |
| dc.title | 探討胚胎神經細胞與老鼠神經瘤細胞內調控細胞存活的蛋白質: F-spondin | zh_TW |
| dc.title | Study of F-spondin protein in the regulation of cell survival of embryonic neuron cells and murine neuroblastoma | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 99-2 | |
| dc.description.degree | 博士 | |
| dc.contributor.coadvisor | 楊淑美(Shu-Mei Liang) | |
| dc.contributor.oralexamcommittee | 梁啟銘(Chi-Ming Liang),陳義雄(Yee-Hsiung Chen),郭呈欽(Cheng-Chin Kuo),楊維元(Wei-Yuan Yang),廖永豐(Yung-Feng Liao) | |
| dc.subject.keyword | 底板反應蛋白,G-四聯體,寡去氧核苷,酸,胚胎神經細胞,YBox 一號 結合蛋白,第六號介白素,p38有絲分裂活化蛋白激酶,神經細胞瘤,神經細胞存活,神經退化疾病,神經疾病, | zh_TW |
| dc.subject.keyword | F-spondin,G-quadruplex,oligodeoxynucleotide,Y box binding protein,interleukin-6,p38 MAPK,embryonic neuron cell,neuroblastoma,neuronal survival,neurodegenerative disease,neurological disorders, | en |
| dc.relation.page | 131 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2011-08-01 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 生化科學研究所 | zh_TW |
| 顯示於系所單位： | 生化科學研究所 | |
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