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http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/81976完整後設資料紀錄
| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 陳漪紋(Yi-Wen Chen) | |
| dc.contributor.author | Ching-Chu Yu | en |
| dc.contributor.author | 虞景筑 | zh_TW |
| dc.date.accessioned | 2022-11-25T05:33:28Z | - |
| dc.date.available | 2026-09-09 | |
| dc.date.copyright | 2021-11-09 | |
| dc.date.issued | 2021 | |
| dc.date.submitted | 2021-09-14 | |
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Functional characterization of extracellular vesicles produced by Bacteroides gingivalis. Infect Immun 55, 111-117, doi:10.1128/iai.55.1.111-117.1987 (1987). 37 Wei, S. et al. Outer membrane vesicles enhance tau phosphorylation and contribute to cognitive impairment. J Cell Physiol 235, 4843-4855, doi:10.1002/jcp.29362 (2020). 38 Dominy, S. S. et al. Porphyromonas gingivalis in Alzheimer’s disease brains: Evidence for disease causation and treatment with small-molecule inhibitors. Science Advances 5, eaau3333, doi:10.1126/sciadv.aau3333 (2019). 39 Ishida, N. et al. Periodontitis induced by bacterial infection exacerbates features of Alzheimer’s disease in transgenic mice. npj Aging and Mechanisms of Disease 3, 15, doi:10.1038/s41514-017-0015-x (2017). 40 Ryder, M. I. Porphyromonas gingivalis and Alzheimer disease: Recent findings and potential therapies. Journal of Periodontology 91, S45-S49, doi:https://doi.org/10.1002/JPER.20-0104 (2020). 41 Haditsch, U. et al. Alzheimer's Disease-Like Neurodegeneration in Porphyromonas gingivalis Infected Neurons with Persistent Expression of Active Gingipains. J Alzheimers Dis 75, 1361-1376, doi:10.3233/jad-200393 (2020). 42 Mantri, C. K. et al. Fimbriae-mediated outer membrane vesicle production and invasion of Porphyromonas gingivalis. Microbiologyopen 4, 53-65, doi:10.1002/mbo3.221 (2015). 43 Seyama, M. et al. Outer membrane vesicles of Porphyromonas gingivalis attenuate insulin sensitivity by delivering gingipains to the liver. Biochim Biophys Acta Mol Basis Dis 1866, 165731, doi:10.1016/j.bbadis.2020.165731 (2020). 44 Ho, M. H., Chen, C. H., Goodwin, J. S., Wang, B. Y. Xie, H. Functional Advantages of Porphyromonas gingivalis Vesicles. PLoS One 10, e0123448, doi:10.1371/journal.pone.0123448 (2015). 45 Veith, P. D. et al. Porphyromonas gingivalis Outer Membrane Vesicles Exclusively Contain Outer Membrane and Periplasmic Proteins and Carry a Cargo Enriched with Virulence Factors. Journal of Proteome Research 13, 2420-2432, doi:10.1021/pr401227e (2014). 46 Lopes, F. M. et al. Comparison between proliferative and neuron-like SH-SY5Y cells as an in vitro model for Parkinson disease studies. Brain Res 1337, 85-94, doi:10.1016/j.brainres.2010.03.102 (2010). 47 Sheets, S. M., Potempa, J., Travis, J., Fletcher, H. M. Casiano, C. A. Gingipains from Porphyromonas gingivalis W83 synergistically disrupt endothelial cell adhesion and can induce caspase-independent apoptosis. Infection and immunity 74, 5667-5678, doi:10.1128/IAI.01140-05 (2006). 48 Ackerman, K., Fiddler, J., Soh, T. Clarke, S. BV-2 Microglial Cells Used in a Model of Neuroinflammation. The FASEB Journal 29, 608.602, doi:https://doi.org/10.1096/fasebj.29.1_supplement.608.2 (2015). 49 Fleetwood, A. J. et al. Metabolic Remodeling, Inflammasome Activation, and Pyroptosis in Macrophages Stimulated by Porphyromonas gingivalis and Its Outer Membrane Vesicles. Front Cell Infect Microbiol 7, 351, doi:10.3389/fcimb.2017.00351 (2017). 50 Kumar, D. K. V. et al. Amyloid-β peptide protects against microbial infection in mouse and worm models of Alzheimer’s disease. Science Translational Medicine 8, 340ra372, doi:10.1126/scitranslmed.aaf1059 (2016). 51 Brothers, H. M., Gosztyla, M. L. Robinson, S. R. The Physiological Roles of Amyloid-β Peptide Hint at New Ways to Treat Alzheimer's Disease. Frontiers in Aging Neuroscience 10, 118 (2018). 52 Jia, L. et al. Pathogenesis of Important Virulence Factors of Porphyromonas gingivalis via Toll-Like Receptors. Frontiers in Cellular and Infection Microbiology 9, 262 (2019). 53 Andrukhov, O., Ertlschweiger, S., Moritz, A., Bantleon, H. P. Rausch-Fan, X. Different effects of P. gingivalis LPS and E. coli LPS on the expression of interleukin-6 in human gingival fibroblasts. Acta Odontol Scand 72, 337-345, doi:10.3109/00016357.2013.834535 (2014). 54 Furuta, N. et al. Porphyromonas gingivalis Outer Membrane Vesicles Enter Human Epithelial Cells via an Endocytic Pathway and Are Sorted to Lysosomal Compartments. Infection and Immunity 77, 4187-4196, doi:10.1128/IAI.00009-09 (2009). 55 Man, S. M. Kanneganti, T.-D. Regulation of inflammasome activation. Immunological reviews 265, 6-21, doi:10.1111/imr.12296 (2015). 56 Elizagaray, M. L. et al. Canonical and Non-canonical Inflammasome Activation by Outer Membrane Vesicles Derived From Bordetella pertussis. Frontiers in Immunology 11, 1879 (2020). | |
| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/81976 | - |
| dc.description.abstract | "阿茲海默症(Alzheimer’s Disease, AD)，是一種神經退化性疾病，約佔失智人口當中的70%。在阿茲海默症患者腦中發現會有大量神經元的死亡。在其病理特徵中，發現有乙型類澱粉蛋白的大量累積，也有微膠細胞(microglial cell)的活化，兩者會直接和間接地對神經元造成傷害。雖然已經對阿茲海默症的病因有些微了解，但確切成因仍有待進一步釐清。先前研究指出，在阿茲海默症病患腦中有發現一種牙周病的主要致病菌-牙齦卟啉單胞菌(Porphyromonas gingivalis)與其毒素(gingipain)。另一研究發現，從AD病患糞便純化出的細菌的外膜囊泡(outer membrane vesicles, OMV)，能夠穿越血腦屏障(blood-brain barrier)，並促使濤蛋白(tau)的磷酸化及損害小鼠的認知能力。因此我們假設P. gingivalis的OMV (Pg-OMVs)也能穿過血腦屏障，促進阿茲海默症的病理現象。 本實驗使用P. gingivalis (ATCC #33277)之菌株用以分離其OMV，並使用穿透式電子顯微鏡確認細菌外膜囊泡的結構。並以神經細胞株SH-SY5Y及N2A和微膠細胞株BV-2作為模型來進行體外試驗(in vitro)。使用不同濃度的OMV，進行細胞形態及細胞活性變化的觀察，並以西方墨點法(Western Blot)檢視神經元之細胞凋亡蛋白與微膠細胞之發炎蛋白的表現情形。 實驗結果顯示，分離出的Pg-OMVs為直徑50-150nm大小的球形結構。隨著Pg-OMVs的濃度上升，神經元皺縮及漂浮的情形越顯著、細胞活性也隨之下降，並導致細胞凋亡蛋白(PARP)的截切；另外，Pg-OMVs也使得微膠細胞的細胞活性下降及誘導其發炎蛋白(iNOS、TNF-alpha)的表現。有趣的是，我們的實驗更發現，原以為會與Pg-OMVs一同對細胞造成傷害加成效果的乙型類澱粉蛋白寡聚體，會專一性地抑制Pg-OMVs對神經元與微膠細胞造成的毒性。 雖然在細胞實驗中有證實Pg-OMVs會導致微膠細胞的發炎以及神經元的死亡，與阿茲海默症病癥相符，但在動物體內Pg-OMVs是否真能成功從牙周組織入侵到腦部，再對腦內神經元造成損傷，以及乙型類澱粉蛋白寡聚體抑制Pg-OMVs毒性的機制為何，仍需進一步進行後續實驗做釐清。" | zh_TW |
| dc.description.provenance | Made available in DSpace on 2022-11-25T05:33:28Z (GMT). No. of bitstreams: 1 U0001-0909202113404200.pdf: 5391233 bytes, checksum: 87e68ac7c5f4b0feb8f2d97044cd7250 (MD5) Previous issue date: 2021 | en |
| dc.description.tableofcontents | 誌謝………………………………………………………………………I 中文摘要………………………………………………………………II ABSTRACT……………………………………………………….… III 目錄…………………………………………………………………VI 圖目錄………………………………………………………………X 表目錄………………………………………………………………XII CHAPTER 1 緒論…………………………………………………..1 CHAPTER 2 文獻回顧……………………………………………..3 2.1 阿茲海默症………………………………………………………………3 2.1.1 乙型類澱粉蛋白…………………………………………………………3 2.1.2 類澱粉梯瀑理論………………………………………………………4 2.1.3 微膠細胞介導的神經元損傷…………………………………………6 2.2 牙齦卟啉單胞菌………………………………………………………7 2.2.1 牙周病…………………………………………………………………7 2.2.2 外膜囊泡………………………………………………………………8 2.3 相互作用………………………………………………………………9 2.3.1 牙齦卟啉單胞菌與阿茲海默症………………………………………9 CHAPTER 3 研究動機與目的……………………………………..10 3.1 研究動機…………………………………………………………………10 3.2 研究目的…………………………………………………………………10 CHAPTER 4 實驗材料、儀器與方法……………………………….11 4.1 實驗材料………………………………………………………………11 4.1.1 細菌培養…………………………………………………………11 4.1.2 細胞培養……………………………………………………………15 4.1.3 外膜囊泡分離………………………………………………………16 4.1.4 西方墨點法…………………………………………………………17 4.1.5 細胞存活率分析…………………………………………………18 4.1.6 牙齦蛋白酶活性測試………………………………………………18 4.1.7 牙齦蛋白酶活性抑制劑 (gingipain inhibitor)………………19 4.1.8 beta-amyloid oligomer與beta-amyloid fibril製備………………………………20 4.1.9 Enzyme-linked immunosorbent assay (ELISA)…………20 4.1.10 其他實驗耗材……………………………………………………21 4.2 實驗儀器……………………………………………………………22 4.3 實驗方法……………………………………………………………24 4.3.1 beta-amyloid oligomer與beta-amyloid fibril製備方法………………………24 4.3.2 細菌培養、保存與外膜囊泡純化方法……………………………25 4.3.3 細菌實驗方法………………………………………………………27 4.3.4 細胞培養與保存方法………………………………………………28 4.3.5 細胞活性測試(MTT reduction assay)…………………………32 4.3.6 西方墨點法(Western blotting)…………………………………33 4.3.7 蛋白質定量-BCA Assay…………………………………………43 4.3.8 酵素結合免疫吸附分析法(ELISA)………………………………44 4.3.9 細胞實驗……………………………………………………………45 4.3.10 統計分析……………………………………………………………46 CHAPTER 5實驗結果………………………………………………..47 5.1 牙齦卟啉單胞菌釋放帶有牙齦蛋白酶之外膜囊泡……………………47 5.2 Pg-OMVs會使神經細胞與微膠細胞之活性下降…………………49 5.3 Pg-OMVs會造成神經細胞與微膠細胞之型態變化………………51 5.4 Pg-OMVs誘導神經細胞走向細胞凋亡……………………………53 5.5 Pg-OMVs帶有的Arg-gingipain(Rgp)是造成SH-SY5Y細胞活性下降的主要原因. ……………………………55 5.6 Pg-OMVs可以活化BV-2產生發炎因子……………………………56 5.7 beta-amyloid oligomer與beta-amyloid fibril製備………………………………………59 5.8 beta-amyloid oligomer可以保護細胞株因Pg-OMVs所造成的毒性…………60 5.8.1 beta-amyloid oligomer可以逆轉Pg-OMVs造成的細胞形態改變……………60 5.8.2 beta-amyloid oligomer可以抑制Pg-OMVs對SH-SY5Y造成的細胞活性下降.62 5.8.3 beta-amyloid oligomer可以抑制Pg-OMVs誘導的微膠細胞活化……………63 5.9 beta-amyloid fibril對於BV-2細胞的影響與beta-amyloid oligomer有不同的結果………66 5.10 beta-amyloid oligomer對於不同細菌與毒素種類造成的發炎反應有不同的影響…69 CHAPTER 6 討論…………………………………………………..75 CHAPTER 7 結論…………………………………………………..82 CHAPTER 8 參考資料……………………………………………..83 | |
| dc.language.iso | zh-TW | |
| dc.subject | 牙周病 | zh_TW |
| dc.subject | 乙型類澱粉蛋白 | zh_TW |
| dc.subject | 外膜囊泡 | zh_TW |
| dc.subject | 牙齦卟啉單胞菌 | zh_TW |
| dc.subject | 阿茲海默症 | zh_TW |
| dc.subject | Alzheimer’s Disease | en |
| dc.subject | Outer membrane vesicle | en |
| dc.subject | Porphyromonas gingivalis | en |
| dc.subject | Beta-amyloid | en |
| dc.subject | Periodontal Disease | en |
| dc.title | 牙齦卟啉單胞菌之外膜囊泡對神經細胞與微膠細胞的影響及阿茲海默症之致病機轉 | zh_TW |
| dc.title | The Effects of Outer Membrane Vesicles Derived from Porphyromonas gingivalis on Neuron and Microglia in the Pathogenesis of Alzheimer’s Disease | en |
| dc.date.schoolyear | 109-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.coadvisor | 李伯訓(Bor-Shiunn Lee) | |
| dc.contributor.oralexamcommittee | 洪千岱(Hsin-Tsai Liu),蕭永基(Chih-Yang Tseng) | |
| dc.subject.keyword | 牙周病,阿茲海默症,牙齦卟啉單胞菌,外膜囊泡,乙型類澱粉蛋白, | zh_TW |
| dc.subject.keyword | Periodontal Disease,Alzheimer’s Disease,Porphyromonas gingivalis,Outer membrane vesicle,Beta-amyloid, | en |
| dc.relation.page | 92 | |
| dc.identifier.doi | 10.6342/NTU202103082 | |
| dc.rights.note | 同意授權(限校園內公開) | |
| dc.date.accepted | 2021-09-14 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 口腔生物科學研究所 | zh_TW |
| dc.date.embargo-lift | 2026-09-09 | - |
| 顯示於系所單位： | 口腔生物科學研究所 | |
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