Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/92284
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dc.contributor許美鈴zh_TW
dc.contributorMeei-Ling Sheuen_US
dc.contributor.author劉珈妤zh_TW
dc.contributor.authorChia Yu Liuen_US
dc.contributor.other生物醫學研究所zh_TW
dc.date2014zh_TW
dc.date.accessioned2015-12-15T05:42:14Z-
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dc.identifier.urihttp://hdl.handle.net/11455/92284-
dc.description.abstract胃癌的全球死亡率高居不下,除了胃癌難以根治,更重要的是術後會有轉移的高風險,因此積極尋找一個新的治療策略顯得至關重要,而在過去的文獻指出CD47在癌細胞逃脫吞噬作用和轉移扮演一個重要角色,且在眾多癌症中已被發現是過度表達,而中和性抗體CD47能有效抑制黑色素瘤、乳癌等腫瘤生長,但在胃癌的角色及機轉尚待釐清。在本篇研究中,我們證明使用中和性抗體治療,標靶分子CD47可以抗腫瘤生長和腹膜轉移。利用西方點墨法和免疫染色法證明在人類胃癌細胞與致癌物質MNNG誘導老鼠腫瘤發生中,發現CD47高度表現。在裸鼠腫瘤異體移植試驗中,經由正子電腦斷層掃描(PET/CT)也清楚驗證CD47抗體有效的減少腫瘤在腹膜中的擴散轉移。更重要的是CD47抗體有效抑制芳香烴受體(AhR)和Snail,且增加上皮細胞指標蛋白,如细胞角蛋白-18(Cytokeratin 18),另外活化內質網壓力(ER stress)指標蛋白Calpain-10。透過免疫沈澱法證明CD47抗體治療提升AhR和Calpain-10交互作用。從電泳遷移率實驗(EMSA)的實驗結果我們更進一步證實CD47抗體有效降低轉錄因子Snail和Cytokeratin 18的轉錄作用並減少兩個蛋白間的交互作用。此外,從共軛焦顯微鏡影像圖也證實CD47抗體治療會增加Calpain-10和Cytokeratin 18。另一方面,由正子電腦斷層掃描(PET/CT)有效證明CD47抗體抑制大腸癌、肺癌、乳癌細胞所引起的腹膜轉移。綜合以上的實驗結果得知,使用CD47抗體治療可以透過活化ER stress和抑制EMT,進而降低胃癌的腫瘤生長和腹膜轉移,顯示CD47抗體可以作為未來治療胃癌及其腹膜轉移的良好策略之一。zh_TW
dc.description.abstractCD47 (Integrin-associated protein, IAP) participates in evade phagocytosis and metastasis but its role in gastric cancer is unclear. We set out to elucidate the expression profile and function of CD47 antibody therapy during gastric antitumor growth and antiperitoneal dissemination effects. Immunoblot and immunohistochemical studies revealed elevated CD47 expression in human gastric cancer cell lines and carcinogen MNNG-induced gastric cancer tissues in animal model. Treated CD47 antibody significantly reduced peritoneal dissemination in a mouse model by positron emission tomography/computed tomography (PET/CT) imaging. Simultaneously, therapeutic antibodies down-regulated AhR, Snail expression in tumor nodules, increased epithelial signatures such as cytokeratin-18 (CK 18), ER stress marker and calpain-10 activation by western blotting. Immuno-precipitation assay, proved adding CD47 therapy promoted AhR/calpain-10 interaction. CD47 therapy efficiently abolished physical interaction and mutual functional between Snail and CK 18 by EMSA. Confocal microscope image demonstrated that CD47 therapy-induced up-regulation of CK 18 translocation was abrogated by calpain-10 blocked. Treated neutralize CD47 antibody efficiently reduced all of peritoneal carcinomatosis such as colon, lung and breast cancer. Taken together, our results suggest that the therapeutic CD47 antibody suppresses both gastric tumor growth and peritoneal dissemination by inducing ER stress and inhibiting EMT.en_US
dc.description.tableofcontents中文摘要 ............................................................ i 英文摘要 ........................................................... ii 目次 .............................................................. iii 圖目次 .............................................................. v 縮寫表 ............................................................. vi 第一章、前言 ........................................................ 1 一、胃癌(Gastric cancer): ....................................................................................... 1 (一)流行病學: .................................................................................................. 1 (二)治療方式: .................................................................................................. 2 二、整合素相關蛋白(Integrin-associated protein,IAP 或 CD47): ......................... 2 三、上皮-間質細胞轉換 (Epithelial mesenchymal transition): ............................ 3 四、細胞角蛋白 18 (Cytokeratin 18): ................................................................... 4 五、芳香烴受體 (Aryl hydrocarbon receptor): ................................................. 4 六、內質網壓力(Endoplasmic Reticulum Stress): ................................................. 4 (一)ER Stress 的分子機轉 ............................................................................... 5 (二)ER Stress 和 Calpain ................................................................................. 6 七、研究方向與動機: ............................................................................................. 6 第二章、 材料與方法 ................................................. 7 一、實驗儀器: ......................................................................................................... 7 二、實驗材料: ......................................................................................................... 7 三、實驗方法: ......................................................................................................... 7 (一)細胞培養(Cell culture) .............................................................................. 7 (二)人類臍帶靜脈內皮細胞初代培養(Primary HUVEC culture) ................. 8 (三)蛋白質萃取(Protein extraction) ................................................................ 8 (四)西方墨點法(Western Blot) ........................................................................ 8 (五)免疫螢光染色法(Immunofluorescence stain)........................................... 9 (六)免疫組織染色(Immunohistochemistry stain) ........................................... 9 (七)免疫沉澱法(Immunoprecipitation) ......................................................... 10 (八)核酸干擾技術(RNA interference) .......................................................... 10 (九)傷口癒合試驗(Wound Healing Assay) ................................................... 10 (十)動物實驗(Animal experiment) ................................................................ 10 (十一)電泳移動率試驗(Electrophoretic Mobility Shift Assay, EMSA)....... 11 (十二)統計 ..................................................................................................... 11 第三章、實驗結果 ................................................... 12 一、在胃癌、致癌物質 MNNG 誘導癌化過程中,高度表現 CD47 的蛋白質量........................................................................................................................ 12 二、以 PET/CT 影像觀察 N/A CD47 抑制胃癌細胞在裸鼠體內生長和腹膜轉移 之情形 ............................................................................................................ 12 三、N/A CD47 減緩上皮間質細胞轉化(Epithelial-mesenchymal transition 簡稱 EMT)過程 ...................................................................................................... 12 四、N/A CD47 在轉錄層次上透過 Snail 及 AhR 有效抑胃癌細胞中 Cytokeratin 18 的轉錄表現 ............................................................................................... 13 五、N/A CD47 誘導內質網壓力的形成且降低 AhR 的表現量,促進胃癌細胞 中的 Calpain 10 與 AhR 的交互作用 ........................................................... 13 六、N/A CD47 會抑制由癌細胞所引起的腹膜轉移 .......................................... 14 七、結論 ................................................................................................................ 14 第四章、討論 ....................................................... 15 第五章、參考文獻 ................................................... 17 附錄表 ............................................................. 34 附圖一 ............................................................. 41 附圖二 ............................................................. 42zh_TW
dc.language.isozh_TWzh_TW
dc.rights同意授權瀏覽/列印電子全文服務,2016-06-23起公開。zh_TW
dc.subject胃癌zh_TW
dc.subjectCD47zh_TW
dc.subjectCalpain-10zh_TW
dc.subjectCytokeratin 18zh_TW
dc.subjectAhRzh_TW
dc.subjectEMTzh_TW
dc.subject腹膜轉移zh_TW
dc.subjectGastric canceren_US
dc.subjectCD47en_US
dc.subjectCalpain-10en_US
dc.subjectCK 18;AhR; EMTen_US
dc.subjectperitoneal disseminationen_US
dc.title標靶分子CD47調控胃癌腹膜轉移機轉之探討zh_TW
dc.titleThe mechanism of targeting CD47 regulates gastric cancer cell peritoneal disseminationen_US
dc.typeThesis and Dissertationen_US
dc.date.paperformatopenaccess2016-06-23zh_TW
dc.date.openaccess2016-06-23-
item.fulltextwith fulltext-
item.languageiso639-1zh_TW-
item.openairetypeThesis and Dissertation-
item.cerifentitytypePublications-
item.openairecristypehttp://purl.org/coar/resource_type/c_18cf-
item.grantfulltextrestricted-
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