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http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/63184完整後設資料紀錄
| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 陳進庭 | |
| dc.contributor.author | Pei-Chi Chiang | en |
| dc.contributor.author | 江珮琪 | zh_TW |
| dc.date.accessioned | 2021-06-16T16:26:54Z | - |
| dc.date.available | 2016-02-01 | |
| dc.date.copyright | 2013-02-01 | |
| dc.date.issued | 2013 | |
| dc.date.submitted | 2013-01-18 | |
| dc.identifier.citation | [1] Hanahan D, Weinberg RA. Hallmarks of cancer: the next generation. Cell 2011; 144(5):646-74.
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/63184 | - |
| dc.description.abstract | 光動力療法 (photodynamic therapy, PDT) 為近年所發展出的臨床癌症治療方法。然而，由於受限於光的穿透能力，深層組織中的癌細胞在治療後仍有機會存活下來。因此，探討在PDT處理後存活下來細胞的生理特徵變化，為重要的研究方向之一。本研究的初步結果顯示，經五胺基酮戊酸光動力療法 (δ-aminolevulinic acid mediated photodynamic therapy, ALA-PDT) 處理後存活下來之細胞，轉移能力將會減弱，且此現象與基質金屬蛋白酶9 (matrix metalloproteinase 9, MMP9) 的表現量下降有關。而在利用微陣列晶片分析的結果中顯示，與腫瘤發展機制可能有關的胞內氯離子通道蛋白4 (chloride intracellular channel 4, CLIC4)，於PDT處理後存活下來之細胞中，表現量明顯較低。且不論是在細胞層級或是以活體動物為模型的研究結果皆顯示，光動力致壓效應將會降低CLIC4及MMP9的表現，藉由提升胞內CLIC4之表現量將可阻礙光動力致壓效應對MMP9及癌轉移能力的影響。此外，當直接調控癌細胞內CLIC4之表現時，MMP9及癌轉移的能力也將受到影響，彼此之間呈正相關。另一方面，在探討光動力致壓效應調控CLIC4之分子機轉的結果中顯示，光動力致壓效應所產生的活性氧化物質 (reactive oxygen species, ROS)，將活化胞內抑癌蛋白p53進而抑制CLIC4之表現。綜合上述結果可得知，光動力致壓效應抑制癌轉移之分子調控機制，可能是透過降低胞內CLIC4之表現，進而使得下游MMP9的表現量受到影響，最終導致抑制癌轉移之生物效應發生。 | zh_TW |
| dc.description.provenance | Made available in DSpace on 2021-06-16T16:26:54Z (GMT). No. of bitstreams: 1 ntu-102-F95b47402-1.pdf: 23511318 bytes, checksum: 7a19099cb397f19da82d4c3ccc4036b9 (MD5) Previous issue date: 2013 | en |
| dc.description.tableofcontents | 中文摘要………………………………...……………………………………………… i
英文摘要…………………………...………………………………………………….. ii 縮寫表 …………………………..........................…………………………………….. iii 第一章、 緒論 1.1 癌細胞之侵襲與轉移 (Invasion and Metastasis)………...…………………….. 1 1.1.1 癌細胞轉移之生理特性變化…………………………………….………... 2 1.1.2 癌細胞轉移之相關因子………………………………………….………... 5 1.2 光動力療法 (Photodynamic therapy, PDT)……………….…………………… 8 1.2.1 光動力療法之應用…………………………………………….…………... 8 1.2.2 光化學作用………………………………………………………………… 9 1.2.3 五胺基酮戊酸光動力療法……………………………………………….. 10 1.2.4 光動力療法對腫瘤治療及癌轉移之影響……………………………….....11 1.3 基質金屬蛋白酶 (Matrix metalloproteinases, MMPs) ……………………….. 13 1.4 胞內氯離子通道蛋白 (Chloride intracellular channel 4, CLIC4)…….……… 14 1.5 p53腫瘤抑制蛋白 (p53 tumor suppressor protein)…………………………... 17 1.6 表觀遺傳修飾 (Epigenetic modification) …………………………………….. 18 1.6.1 組蛋白修飾作用 (Histone modification)………………………………….18 1.6.2 DNA甲基化修飾作用 (DNA methylation)………………………………19 1.6.3 DNA甲基轉移酶 (DNA methyltransferase, DNMT)………….…………20 第二章、研究動機與目的…………………………………………………………21 第三章、實驗架構………………………………………………………………….23 第四章、材料與方法 4.1 細胞株 (Cell lines)………………………………………………………………. 24 4.2 藥品 (Chemicals)………………………………………………………………… 24 4.3 細胞培養 (Cell culture)………………………………………………………….. 26 4.4 光動力處理 (Photodynamic therapy, PDT)……………………………………... 27 4.5 細胞存活率分析：粒線體去氫酶活性分析 (MTT assay)………………………29 4.6 細胞群落形成之分析 (Colony formation assay)……………………………….. 29 4.7 明膠蛋白酶活性電泳分析法 (Gelatin Zymography)…………………………... 30 4.8 挑選單一細胞群落 (Single colony selection)…………………………………... 31 4.9 細胞移動能力分析 (Scratch wound healing assay)…………………………….. 31 4.10 細胞侵襲能力分析 (Matrigel invasion assay)…………………………………... 32 4.11 流式細胞儀測定細胞週期 (Flow Cytometry: PI staining)……………………... 32 4.12 胞內Protoporphyrin IX (PpIX) 含量分析……………………………………….33 4.13 粒線體含量分析：檸檬酸合成酶活性分析 (Citrate synthase activity)…………34 4.14 粒線體膜電位分析 (Mitochondrial transmembrane potential)…………………. 35 4.15 穿透式電子顯微鏡 (Transmission electron microscopy)……………………...... .36 4.16 細胞骨架之免疫螢光染色 (Immunofluorescence, IFC)………………………... 36 4.17 西方墨點法 (Western blot)……………………………………………………… 37 4.18 反轉錄聚合酶鏈鎖反應 (RT-PCR)……………………………………………... 39 4.19 cDNA微陣列分析 (cDNA microarray)………………………………………… 42 4.20 細胞轉染 (Plasmid or siRNA Transfection)…………………………………….. 42 4.21 DNA甲基轉移酶酵素活性測試 (DNMTs activity)……………………………. 44 4.22 亞硫酸氫鹽-基因組測序法 (Bisulfite genomic sequencing, BGS) ……………..45 4.23 統計方法 (Statistical analysis)…………………………………………………. 47 第五章、結果 5.1 光動力致壓效應對細胞生理特徵之影響………………………………………. 48 5.1.1 ALA-PDT光動力處理對細胞存活率及形態之影響 5.1.2 ALA-PDT-derived變異株細胞的形態及移動能力之變化 5.2 光動力致壓效應與基質金屬蛋白酶 (MMPs) 之關聯性……………………... 50 5.2.1 ALA-PDT-derived變異株細胞的MMPs基因表現及蛋白酶活性變化 5.2.2 ALA-PDT光動力處理對MMP9基因表現及蛋白酶活性之影響 5.3 光動力致壓效應致使轉移能力減弱的單一形式細胞株之特性分析…………. 52 5.3.1 移動能力 (migration ability) 5.3.2 生長週期 (cell cycle) 5.3.3 細胞骨架排列情形 (cytoskeleton arrangement) 5.3.4 粒線體功能 (mitochondrial function) 5.3.5 基因表現變化 (cDNA microarray analysis) 5.4 光動力致壓效應對胞內氯離子通道蛋白 (CLIC4) 之影響………………….. 59 5.4.1 ALA-PDT光動力處理對CLIC4基因表現之影響 5.4.2 ALA-PDT對PDT-derived變異株、Clone A03及A06細胞存活率之影響 5.4.3 ALA-PDT對PDT-derived變異株、Clone A03及A06細胞基因表現影響 5.4.4 Ce6-PDT光動力處理對CLIC4及MMP9基因表現之影響 5.4.5 於活體動物研究中 (In vivo study)，Ce6-PDT對腫瘤細胞之CLIC4及MMP9基因表現的影響 5.5 CLIC4及MMP9與光動力致壓效應抑制癌轉移之關聯……………………… 64 5.5.1 提升Clone A06細胞之CLIC4表現對MMP9及轉移能力的影響 5.5.2 提升胞內CLIC4之表現對ALA-PDT抑制MMP9及轉移能力的影響 5.6 光動力致壓效應調控CLIC4之分子作用機轉…………………………………67 5.6.1 組蛋白去乙醯化酶抑制劑 (HDACi) 對ALA-PDT調控CLIC4之影響 5.6.2 DNA甲基轉移酶抑制劑 (DNMTi) 對ALA-PDT調控CLIC4之影響 5.6.3 ALA-PDT光動力處理對DNMT活性及基因表現之影響 5.6.4 抑制胞內DNMT1之表現對ALA-PDT調控CLIC4的影響 5.6.5 ALA-PDT光動力處理對CLIC4啟動子區域之甲基化程度的影響 5.6.6 抗氧化劑 (ROS scavenger) 對ALA-PDT調控DNMT1及CLIC4之影響 5.6.7 抑制胞內p53之表現對ALA-PDT調控DNMT1及CLIC4的影響 5.7 CLIC4對癌細胞轉移能力之影響及相關分子作用機轉………………………. 75 5.7.1 抑制invasive癌細胞之CLIC4表現對MMP9及轉移能力的影響 5.7.2 提升non-invasive癌細胞之CLIC4表現對MMP9及轉移能力的影響 5.7.3 調控胞內CLIC4之表現對細胞貼附因子E-cadherin的影響 5.7.4 提升胞內CLIC4之表現對核內p-Smad2 & 3表現的影響 5.7.5 提升胞內CLIC4之表現對NF-κB入核能力的影響 第六章、討論 6.1 CLIC4及MMP9與光動力致壓效應抑制癌轉移之關聯性……………………. 80 6.2 光動力致壓效應誘發細胞死亡的過程與影響基因表現之關聯性……………. 82 6.3 光動力致壓效應對細胞生物效應之影響………………………………………. 83 6.4 CLIC4之相關調控機制…………………………………………………………. 86 6.5 CLIC4與腫瘤發展及癌轉移之關聯………………………………………….…. 89 6.6 CLIC4調控MMP9之分子相關機制……………………………………………. 92 第七章、結論………………………………………………………………………. 93 第八章、未來研究方向……………………………………………………….…. 95 參考文獻………………………………………………………………………...…. 189 | |
| dc.language.iso | zh-TW | |
| dc.subject | 胞內氯離子通道蛋白4 | zh_TW |
| dc.subject | 基質金屬蛋白酶 | zh_TW |
| dc.subject | 光動力效應 | zh_TW |
| dc.subject | 癌轉移 | zh_TW |
| dc.subject | 去氧核醣核酸甲基化 | zh_TW |
| dc.subject | CLIC4 | en |
| dc.subject | MMP9 | en |
| dc.subject | Photodynamic therapy | en |
| dc.subject | Metastasis | en |
| dc.subject | DNA methylation | en |
| dc.title | 探討CLIC4於光動力致氧化壓力下之調控機制及其對細胞生物效應之影響 | zh_TW |
| dc.title | Molecular Mechanisms Involved in PDT-induced Reduction of CLIC4 and its Subsequent Biological Responses | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 101-1 | |
| dc.description.degree | 博士 | |
| dc.contributor.oralexamcommittee | 黃慶璨,張麗冠,蔡翠敏,楊維中 | |
| dc.subject.keyword | 胞內氯離子通道蛋白4,基質金屬蛋白酶,9,光動力效應,癌轉移,去氧核醣核酸甲基化, | zh_TW |
| dc.subject.keyword | CLIC4,MMP9,Photodynamic therapy,Metastasis,DNA methylation, | en |
| dc.relation.page | 209 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2013-01-18 | |
| 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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