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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
---|---|---|
dc.contributor.advisor | 黃楓婷(Feng-Ting Huang) | |
dc.contributor.author | Yen-Hong Lin | en |
dc.contributor.author | 林彥霐 | zh_TW |
dc.date.accessioned | 2021-06-08T03:55:46Z | - |
dc.date.copyright | 2018-08-16 | |
dc.date.issued | 2018 | |
dc.date.submitted | 2018-08-15 | |
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Diabetes, 63(2), 514-525. doi:10.2337/db13-1106 Zielinski, M. R., Muenchow, M., Wallig, M. A., Horn, P. L., & Woods, J. A. (2004). Exercise delays allogeneic tumor growth and reduces intratumoral inflammation and vascularization. Journal of applied physiology, 96(6), 2249-2256. Zou, Z., Zeng, F., Xu, W., Wang, C., Ke, Z., Wang, Q. J., & Deng, F. (2012). PKD2 and PKD3 promote prostate cancer cell invasion via uPA by shifting balance between NF-κB and HDAC1. J Cell Sci, jcs. 106542. 李萱葒. (2015). Irisin對人類神經膠質瘤細胞株U-87 MG之影響. 臺灣大學. Available from Airiti AiritiLibrary database. (2015年) 張育瑄. (2017). 探討Irisin對人類神經膠質瘤細胞株U-87 MG之機制影響. 臺灣大學. Available from Airiti AiritiLibrary database. (2017年) | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/21972 | - |
dc.description.abstract | 運動能夠降低罹患癌症的風險與延緩癌症的進程。運動會誘導肌肉分泌激素,這些激素統稱為肌肉激素。Irisin 也是一種由運動誘導分泌的肌肉激素,它能夠促進白色脂肪細胞分化為類棕色脂肪細胞,此過程稱為褐化作用,且研究指出醣基化能夠提升irisin在褐化作用上的效果。本實驗室先前的研究發現,irisin可以減緩人類神經膠質瘤細胞株U-87 MG侵襲與增生的速度。我們推測具有醣基化的irisin對人類神經膠質瘤細胞株的影響會更加顯著。我們利用酵母菌系統表現人類irisin蛋白質,並探討其對人類神經膠質瘤細胞株的影響。我們的研究顯示,irisin能夠減緩細胞增生但不影響細胞凋亡。另外我們發現酵母菌所生產具有醣基化的irisin,在抑制細胞增生的效果比大腸桿菌所生產沒有醣基化的irisin來的好,這指出醣基化對irisin的生物功能有其重要性。Irisin 透過造成細胞週期停滯減緩細胞增生的速度,包含在irisin處理48小時的G0停滯及irisin處理72小時的G2停滯。另外irisin能夠促進CDK抑制蛋白p21Waf1/Cip1的表現量以及促進cdc2的磷酸化。p21 會結合CDK-cyclin複合體使其失活,並造成細胞週期停滯。Cdc2 負責調控G2/M檢查點,cdc2能透過去磷酸化被活化,並促進細胞通過G2/M檢查點。因此,irisin處理的細胞當中磷酸化的cdc2比例增加也是造成G2停滯的原因之一。除此之外,我們的研究發現,irisin能夠抑制細胞的遷移與侵襲能力。我們也觀察到irisin能夠使細胞當中SerpinB2以及TFPI-2的表現量增加。不過irisin是否是透過促進SerpinB2以及TFPI-2的表現來抑制細胞遷移與侵襲仍然需要更進一步的研究。Irisin是如何促進p21Waf1/Cip1的表現、以及如何造成磷酸化的cdc2比例上升也仍須進一步的實驗研究。儘管irisin對癌症的影響在機制上還需要進一步釐清,我們認為irisin能夠延緩癌症的進程,並且能夠成為預防癌症的新手段。 | zh_TW |
dc.description.abstract | Studies indicate that exercise can reduce the risk of cancer and delay cancer progression. Exercise induces the release of humoral factors from muscle cells, which were called myokines. Irisin is a myokine that can induce browning of adipocytes, and glycosylation could enhance its function in browning. Previous studies of our laboratory found that irisin can reduce cell invasion and proliferation of a human glioblastoma cell line, U-87 MG. We proposed that glycosylated irisin could have higher impact on glioblastoma. Hence, in the thesis, we produced recombinant human irisin protein in Pichia pastoris, and investigated its effects on human glioblastoma cell lines. Our studies showed that irisin reduced cell proliferation but not cell apoptosis. We also found that glycosylated irisin had a better effect on reducing cell proliferation than nonglycosylated irisin from bacteria, indicating that glycosylation has its importance in the biological function of irisin on cancer cells. Irisin reduced cell proliferation by causing cell cycle arrest, including G0 and G2 arrest upon 48 and 72 hours of irisin treatment, respectively. Moreover, irisin induced expression of CDK inhibitor p21Waf1/Cip1 and phosphorylation of cdc2, a cyclin-dependent kinase. p21 binds and inactivates CDK-cyclin complexes and causes cell cycle arrest. Cdc2 regulates the G2/M checkpoint and dephosphorylation activates cdc2 to progress cells through the G2/M checkpoint. Therefore, the ratio of phosphorylated cdc2 increased in irisin treated cells attributed to the G2 arrest of the cell cycle. Furthermore, we found that irisin inhibited cell migration and invasion of U-87 MG cells. Protein levels of SerpinB2 and TFPI-2 were also up-regulated by irisin. However, whether irisin inhibits migration and invasion by inducing SerpinB2 and TFPI-2 still needs further investigation. The mechanism of how irisin induces p21Waf1/Cip1 expression and cause phosphorylation of cdc2 also needs further experiments to find out. Nonetheless, we propose that irisin could delay cancer progression and might be a novel strategy for cancer prevention. | en |
dc.description.provenance | Made available in DSpace on 2021-06-08T03:55:46Z (GMT). No. of bitstreams: 1 ntu-107-R05b22020-1.pdf: 3573967 bytes, checksum: 7b286db669095131bc0e51038f0ea655 (MD5) Previous issue date: 2018 | en |
dc.description.tableofcontents | 謝辭 i
中文摘要 ii Abstract iii Abbreviations v Table of contents vi Chapter 1 Introduction 1 1.1 Exercise, myokine and cancer 1 1.2 Irisin 2 1.2.1 Characteristics and function of irisin 2 1.2.2 Glycosylation of irisin 2 1.2.3 The effect of irisin on cancer 3 1.3 Glioblastoma 4 1.4 Protease inhibitor and tumor invasion 5 1.5 Cell cycle regulators 5 1.6 Research purpose 7 Chapter 2 Material and methods 8 2.1 Recombinant protein preparation 8 2.1.1 Cloning of irisin gene into pPICZα A vector 8 2.1.2 Transformation of pPICZα A-irisin into Pichia pastoris strain X-33 8 2.1.3 Protein induction 9 2.1.4 Protein purification and condensation 9 2.2 PNGaseF treatment 10 2.3 Native-PAGE 10 2.4 Gel filtration 11 2.5 Cell culture 11 2.5.1 Uppsala 87 Malignant Glioma (U-87 MG) 11 2.5.2 T98G cell line 11 2.5.3 LN-18 cell line 11 2.6 Cell migration assay 11 2.7 Cell invasion assay 12 2.8 Cell proliferation analysis 12 2.9 Cell cycle analysis 13 2.10 Cell apoptosis analysis 13 2.11 Cellular protein extraction 14 2.12 Electrophoresis and western blotting 14 2.13 FITC-protein binding assay 15 2.13.1 Labeling FITC to irisin 15 2.13.2 Binding of FITC-labeled irisin to U-87 MG cells 15 Chapter 3 Results 16 3.1 Irisin preparation 16 3.1.1 Construction of recombinant plasmid DNA containing irisin gene and transformation into Pichia pastoris strain X-33 16 3.1.2 Recombinant irisin expression and purification from Pichia pastoris 16 3.1.3 PNGase F treatment of irisin 17 3.1.4 Glycosylation causes polymerization of irisin 17 3.2 Irisin inhibited cell migration of U-87 MG cells 18 3.3 Irisin inhibited cell invasion of U-87 MG cells 18 3.4 Irisin reduced proliferation of glioblastoma cells 19 3.5 Irisin affected cell cycle progression of U-87 MG cell 20 3.5.1 Irisin affected cell distribution of the cell cycle 20 3.5.2 Irisin induced p21Waf1/Cip1 expression in a p53-independent manner and causes cell cycle regulation 21 3.5.3 Irisin increased the protein level of p-cdc2 21 3.6 Irisin bound the cell surface of U-87 MG 22 Chapter 4 Discussion 23 4.1 The effect of glycosylation on irisin function 23 4.2 Irisin caused arrest in cell cycle checkpoints 23 4.3 The effect of irisin on cell migration and invasion 24 Chapter 5 Summary and future prospects 26 Chapter 6 Reference lists 27 Figures 31 Figure 1. Construction of the pPICZα A-irisin plasmid 32 Figure 2. Recombinant irisin expression test and purification 34 Figure 3. PNGase F treatment of irisin 35 Figure 4. Molecular weight determination of glycosylated irisin by gel filtration 38 Figure 5. Irisin inhibited cell migration of U-87 MG 39 Figure 6. Irisin inhibited cell invasion of U-87 MG and increased TFPI-2 and SerpinB2 protein expression 42 Figure 7. Irisin reduced cell proliferation of glioblastoma cells 44 Figure 8. Irisin did not affect cell apoptosis of U-87 MG 45 Figure 9. Irisin affected cell cycle distribution of U-87 MG cells 50 Figure 10. Protein levels of p53 and p21Waf1/Cip1 in irisin treated cells 51 Figure 11. Irisin increased protein level of p-cdc2 53 Figure 12. Irisin bound to the cell surface of U-87 MG and entered into cells 55 Appendixes 56 1. Antibodies 57 | |
dc.language.iso | en | |
dc.title | 探討酵母菌系統表現之Irisin對人類神經膠質瘤細胞株之影響與機制探討 | zh_TW |
dc.title | Study the Effect and the Mechanism of Yeast-produced Irisin on Human Glioblastoma Cell Lines | en |
dc.type | Thesis | |
dc.date.schoolyear | 106-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 周綠蘋,黃兆祺,陳韻如,廖憶純 | |
dc.subject.keyword | irisin,醣基化,細胞增生,細胞遷移,細胞侵襲,細胞週期停滯, | zh_TW |
dc.subject.keyword | irisin,glycosylation,cell proliferation,cell migration,cell invasion,cell cycle arrest, | en |
dc.relation.page | 57 | |
dc.identifier.doi | 10.6342/NTU201803290 | |
dc.rights.note | 未授權 | |
dc.date.accepted | 2018-08-15 | |
dc.contributor.author-college | 生命科學院 | zh_TW |
dc.contributor.author-dept | 生化科技學系 | zh_TW |
顯示於系所單位: | 生化科技學系 |
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