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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 林俊立,賴凌平 | |
| dc.contributor.author | Yuan-Hung Liu | en |
| dc.contributor.author | 劉芫宏 | zh_TW |
| dc.date.accessioned | 2021-06-08T02:39:54Z | - |
| dc.date.copyright | 2018-08-01 | |
| dc.date.issued | 2018 | |
| dc.date.submitted | 2018-06-03 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/20089 | - |
| dc.description.abstract | 心肌梗塞與心臟衰竭的盛行率及死亡率一直高居不下，其中一個最重要的原因是心肌細胞的數量減少到心臟無法代償的程度，目前治療方式並無法有效改善此現象，而心臟再生被認為是最有潛力的治療方法之一。心臟再生在哺乳動物是否存在一直是個問號，最近的研究發現出生後哺乳動物的心肌細胞有再生的現象，但再生細胞的起源並不清楚。目前並沒有直接的證據顯示內源性心臟先驅細胞在心肌受損後可分化為心肌細胞來修補受損的心臟。在胚胎發育期，Nkx2.5+心臟先驅細胞可分裂並分化為心臟內的心肌細胞、血管平滑肌細胞及部分內皮細胞而架構成心臟。
我們的研究的目的是 (1)證實心臟先驅細胞存在出生後哺乳動物心臟且可分化為心肌細胞，(2)證明出生後哺乳動物心臟先驅細胞具再生心臟的能力，(3)確認心臟先驅細胞的來源，(4)探討哺乳動物心臟再生的機轉，(5)應用細胞移植增進心臟再生及修補。我們可以進一步將心臟再生應用在人類受損心臟的修補。 我們發現在心肌受損後心外層的心肌生成基因會被活化，表現類似胚胎心臟發育過程的基因變化，我們進一步證實出生後Nkx2.5+細胞可分化為心肌細胞，Nkx2.5 enhancer-eGFP基因轉殖鼠在心肌梗塞後Nkx2.5 enhancer-eGFP+細胞顯著增加。我們並發現Nkx2.5 enhancer-eGFP+多位於心外膜下 (subepicardium)且表現心臟中胚層標記GATA4和心臟先驅細胞標記Nkx2.5，但不表現血球細胞、纖維母細胞、血球幹細胞及心肌細胞等標記。微陣列分析Nkx2.5 enhancer-eGFP+細胞發現心臟發育相關基因表現顯著增加。我們進一步用雙基因轉殖鼠Inducible Nkx2.5 enhancer Cre/R26R-lacZ作lineage tracing，結果發現Nkx2.5+及其衍生細胞在心臟受損後被活化，由心臟心包膜下逐步移動至心肌層內並分化為心肌細胞。當去除心肌梗塞後的心臟先驅細胞，我們發現心臟功能惡化，此結果提供內源性心臟再生功能性證據。我們更發現使用抗發炎藥物會抑制心臟先驅細胞的活化並減少心肌細胞再生，這顯示心肌梗塞後發炎反應可促進心肌再生。 為探討出生後心臟先驅細胞的來源，我們用三基因轉殖鼠不同的lineage Cre/Nkx2.5 enhancer-eGFP/R26R-LacZ，在心肌梗塞後分析Nkx2.5 enhancer-eGFP+細胞的源頭，發現出後Nkx2.5 enhancer-eGFP+細胞源自胚胎發育期的心外膜細胞 (embryonic epicardial cells)。 我們的研究提供心臟先驅細胞在心肌受損後可分化為心肌細胞來修補受損的心臟的直接證據並證實其來自胚胎發育期的心外膜細胞，此研究有助於了解調節心臟再生的影響因子及機轉，這些研究結果可望在心臟修補及再生治療提供重要的貢獻，未來若能應用於心肌梗塞及心臟衰竭的病人的治療，將可大幅改善病人的預後。 為研究細胞治療是可促進心臟再生及修補，我們使用人類胎盤幹細胞 (human placenta-derived multipotent cells, hPDMCs)作研究。人類胎盤幹細胞已知具三胚層細胞分化的能力，此外它的取得無倫理上的爭議、容易分離、低排斥及具免疫調控能力等特性使它成為非常有潛力的細胞治療方式。我們發現它可表現心臟發育的轉錄因子GATA4及Hand2，我們假設它可透過心肌分化潛力而具心臟修復的能力。 我們證實人類胎盤幹細胞可改善心臟功能，其機制是透過心肌細胞心生、促進血管生成功能及抑制心肌細胞凋亡 (apoptosis)，我們可進一步將這無倫理上的爭議、容易分離、低排斥及具免疫調控能力的細胞用於治療嚴重心血管疾病的病患以改善他們的預後。 | zh_TW |
| dc.description.abstract | Recent studies report that postnatal mammalian hearts undergo cardiomyocyte refreshment; however, evidence is lacking for the cell origin of the cells involved in postnatal cardiomyogenesis.
We first confirmed that myocardial infarction (MI) triggers the expression of embryonic cardiogenesis genes in the external aspects of the heart. We further documented that Nkx2.5 enhancer-eGFP (Nkx2.5 enh-eGFP) cells exist in the postnatal Nkx2.5 enh-eGFP transgenic mice and would differentiate into striated cardiomyocytes in vitro. We created coronary artery ligation on the Nkx2.5 enh-eGFP transgenic mice. The number of Nkx2.5 enh-eGFP cells increased following MI. The Nkx2.5 enh-eGFP cells resided mostly in the subepicardium and expressed precardiac mesoderm marker GATA4 and cardiac precursor marker Nkx2.5. eGFP signaling was not expressed in mature cardiomyocytes, hematopoietic cells or fibroblasts. Transcriptomic analysis of activated Nkx2.5 enh-eGFP cells showed heart development genes up-regulated remarkably and significantly. Using inducible Nkx2.5 enhancer-Cre/ROSA26 reporter double transgenic mice to lineage trace the fate of activated Nkx2.5 enh-eGFP cells, we documented that the activated Nkx2.5 enh-eGFP cells proliferate and differentiate into mature cardiomyocyte in vivo. To trace the developmental origin of the activated Nkx2.5 enh-eGFP cells, we created different lineage-Cre/Nkx2.5 enh-eGFP/ROSA26 reporter triple transgenic mice. Post-MI Nkx2.5 enh-eGFP+ cells originated from the embryonic epicardial cells, not from the pre-existing cardiomyocytes, endothelial cells, cardiac neural crest cells, or perinatal/postnatal epicardial cells. Together, this study confirmed that cardiac lineage-specific progenitor cells, which originate from embryonic epicardium-derived cells, contribute to postnatal mammalian cardiomyogenesis. The discovery of a cardiomyogenic cell population in the postnatal heart enables future cell therapy for cardiac regeneration. Human placenta-derived multipotent cells (hPDMCs) have the capacity of multilineage differentiation into cells with ectodermal, mesodermal, and endodermal phenotypes. These hPDMCs can be obtained without ethical concerns or the need for invasive procedures. Also, their relative ease of isolation and immunosuppressive properties make these multilineage cells very good candidates for cell therapy to treat damaged organs. We confirmed that hPDMCs can modulate cardiac injury in small and large animal models of myocardial injury and elucidate the mechanisms involved. We found that hPDMCs can undergo in vitro cardiomyogenic differentiation when cocultured with mouse neonatal cardiomyocytes. Moreover, hPDMCs exert strong proangiogenic responses in vitro toward human endothelial cells mediated by secretion of hepatocyte growth factor, growth-regulated oncogene-α, and interleukin-8. To test the in vivo relevance of these results, small and large animal models of acute myocardil injury were induced in mice and minipigs. Transplantation of hPDMCs into the animal heart post-acute MI induction improved left ventricular function, with significantly enhanced vascularity, cardiomyogenic differentiation, and antiapoptotic effects. Our study offers mechanistic insights and preclinical evidence on using hPDMCs as a therapeutic strategy to treat severe cardiovascular diseases. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-08T02:39:54Z (GMT). No. of bitstreams: 1 ntu-107-D92421005-1.pdf: 5980560 bytes, checksum: 11012ddf3993bbb5bb7f637c7d5ec494 (MD5) Previous issue date: 2018 | en |
| dc.description.tableofcontents | 口試委員會審定書………………………………………………………….……..….1
誌謝……………………………………………………………………….…….….….2 中文摘要……………………………………………………………………..…….….3 英文摘要………………………………………………………….………….…….….5 博士論文內容 1. 緒論 (Introduction) ……………………………………….…………………….10 1.1 心臟外的先驅細胞在心臟修復的角色…………………………….……..12 1.2 心臟再生的證據…………………………………………………….. ……15 1.3 胚胎心臟發育中的心臟先驅細胞 ……………………………….……….16 1.4 出生後的心臟先驅細胞………………………………………….………..19 1.5 心臟先驅細胞在治療心臟血管疾病的應用……………………………...21 1.6 Nkx2.5……………………………………………………………………..23 1.7 人類胎盤幹細胞…………………………………………………………...24 1.8 研究假說、目的、重要性及說明…………………………………………25 2. 研究方法與材料 (Methods and Materials)……………………………………26 2.1 心臟先驅細胞在心臟再生角色…………………………………………...26 2.2 人類胎盤幹細胞促進心臟再生及修補…………………………………...33 3. 結果 (Results)………………………………………………………………….39 3.1 心臟先驅細胞在心臟再生角色…………………………………………...39 3.1.1心臟受損後啟動心臟新生的基因表現…………………………….39 3.1.2 Nkx2.5 enhancer-eGFP基因轉殖小鼠綠色螢光細胞的表現……..41 3.1.3出生後心臟特異性先驅細胞具分化為心肌細胞的能力………..42 3.1.4 Nkx2.5 enhancer表現細胞在心臟受損後顯著增生………………43 3.1.5 出生後Nkx2.5 enhancer表現細胞在心臟的分布………………..46 3.1.6 出生後Nkx2.5 enhancer表現細胞表現心臟特異性先驅細胞的標誌但不表現成熟心肌細胞的標誌……………………………………………..47 3.1.7 出生後Nkx2.5 enhancer表現細胞的特性………………………..49 3.1.8 出生後Nkx2.5 enhancer表現細胞並非心肌細胞………………...51 3.1.9 出生後Nkx2.5 enhancer表現細胞的功能性分析………………..53 3.1.10 心肌梗塞後Nkx2.5 enhancer表現細胞於體內分化為心肌細胞-- 譜系追踪………………………………………………………………………..55 3.1.11 新生小鼠出生後的Nkx2.5 enhancer表現細胞的譜系追踪……59 3.1.12 心肌梗塞促進Nkx2.5 enhancer表現細胞表達心臟生成的相關基因 ………………………………………………………………………………61 3.1.13 功能性心臟再生………………………………………………...64 3.1.14 發炎反應會活化心臟先驅細胞進行心臟修補………………….67 3.1.15 出生後心臟先驅細胞的發育源頭……………………………….70 3.1.16 胚胎期的心外膜:出生後心臟先驅細胞的發育源頭……………75 3.2 人類胎盤幹細胞促進心臟再生及修補…………………………………...79 3.2.1 人類胎盤幹細胞表現心肌生成基因………………………………79 3.2.2 體外證據顯示hPDMCs可分化成心肌細胞………………………82 3.2.3 體外證據顯示hPDMCs促進血管生成……………………………82 3.2.4 hPDMCs心臟保護作用的分子機制……………………………….83 3.2.5 hPDMCs在小鼠心肌梗塞調節心臟損傷………………………….85 3.2.6 hPDMCs移植後小鼠損傷心臟的血管增加……………………….87 3.2.7 hPDMCs在豬心肌梗塞減少心臟損傷…………………………….89 3.2.8 hPDMCs促進豬心肌梗塞後心臟的血管生成…………………….91 3.2.9 hPDMCs在豬心內可分化為心肌細胞………………….…………94 3.2.10 hPDMCs抑制心肌細胞凋亡……………………………………...96 4. 討論……………………………………………………………………………..98 5. 展望……………………………………………………………………………111 6. 論文英文簡述…………………………………………………………………115 7. 參考文獻………………………………………………………………………170 附錄：博士班修業期間所發表之相關論文清冊…………………………………187 | |
| dc.language.iso | zh-TW | |
| dc.title | 出生後心臟的再生與細胞移植 | zh_TW |
| dc.title | Cardiac Regeneration in the Postnatal Heart and Cell-Based Transplantation | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 106-2 | |
| dc.description.degree | 博士 | |
| dc.contributor.oralexamcommittee | 楊偉勛,蘇銘嘉,陳信孚,林幸榮,顏伶汝 | |
| dc.subject.keyword | 心臟再生,心臟先驅細胞,Nkx2.5,心肌梗塞,心臟衰竭,細胞移植,人類胎盤幹細胞, | zh_TW |
| dc.subject.keyword | cardiac regeneration,cardiac progenitor cells,Nkx2.5,myocardial infarction,heart failure,cell transplantation,human placenta-derived multipotent cells (hPDMCs), | en |
| dc.relation.page | 187 | |
| dc.identifier.doi | 10.6342/NTU201800870 | |
| dc.rights.note | 未授權 | |
| dc.date.accepted | 2018-06-04 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 臨床醫學研究所 | zh_TW |
| 顯示於系所單位： | 臨床醫學研究所 | |
文件中的檔案：
| 檔案 | 大小 | 格式 | |
|---|---|---|---|
| ntu-107-1.pdf 未授權公開取用 | 5.84 MB | Adobe PDF |
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