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| ???org.dspace.app.webui.jsptag.ItemTag.dcfield??? | Value | Language |
|---|---|---|
| dc.contributor.advisor | 林劭品(Shau-Ping Lin) | |
| dc.contributor.author | Ching-Yu Chuang | en |
| dc.contributor.author | 莊靜玉 | zh_TW |
| dc.date.accessioned | 2021-06-17T00:11:02Z | - |
| dc.date.available | 2012-07-27 | |
| dc.date.copyright | 2012-07-27 | |
| dc.date.issued | 2012 | |
| dc.date.submitted | 2012-07-13 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/65751 | - |
| dc.description.abstract | 在研究生殖細胞的發育過程中，人類胚胎幹細胞已被廣泛的證實為一個相當重要的平台。由於取得人類胚胎發育中生殖細胞之樣本極為困難，為了能更進一步的瞭解生殖細胞在分子、訊息和後生遺傳學上的機轉，建立一個能有效的從人類胚胎幹細胞中分離或純化生殖細胞的系統，將會是一個相當關鍵的步驟。在本論文所闡述的實驗中，我們建立了一個能由正在多方分化之人類胚胎幹細胞群中，有效純化生殖細胞的系統，並藉此發現體外誘導生殖細胞分化的重要訊息分子。首先，我們以OCT4-EGFP做為生殖細胞的篩選，藉由此種方式，我們發現可以成功的提升具有減數分裂能力的類生殖細胞之比率。再則，我們發現EpCAM這個表達於全能幹細胞的表面蛋白分子，也同時表現在人類胎兒的性腺上，並可以被應用為體外分化生殖細胞的純化因子。結合OCT4和EpCAM進行双重篩選，則能更有效率的純化出具有減數分裂能力的類生殖細胞。另外，以 OCT4/EpCAM純化出的類生殖細胞之比例做為判讀的指標，我們證實BMP4/pSMAD1/5/8和WNT3A/β-CATENIN具有協同促進作用，能促進人類胚胎幹細胞朝向分化生殖細胞的過程。因此結合BMP4/WNT3A的誘導和OCT4/EpCAM的篩選，我們更進一步明顯的提升具有減數分裂能力的類生殖細胞數量。為了驗證以上述分離的類生殖細胞，在體內的環境中是否能更進一步的發育為成熟的配子，我們將這些BMP4/WNT3A誘導的OCT4+/EpCAM+細胞跟新生老鼠的卵巢細胞混合後，打入具有免疫缺陷之老鼠體內。我們發現這些細胞在體內的環境中，形成更多的的生殖細胞群。因此我們認為我們所建立之純化生殖細胞的詳細步驟和系統，提供了一個非常有用的平台。不但能做為研究生殖細胞發育機轉的有用工具，同時更希望對於未來人類生殖和再生醫學能有更進一步的助益。 | zh_TW |
| dc.description.abstract | Human embryonic stem cells (hESCs) have been proven to be an important platform for studying the molecular, signaling and epigenetic processes of germ cell development. The establishment of an effective germ cell selection/enrichment system from differentiating hESCs is crucial for achieving these goals. In the studies we described in this thesis, we developed a germ cell-enriching system that enables us to identify signaling factors involved in germ cell-fate induction from differentiating hESCs in vitro. Firstly, we demonstrated that selection through an OCT4-EGFP reporter system can successfully increase the percentage of meiotic-competent, germ cell-like cells from spontaneously differentiating hESCs. Furthermore, we showed that the pluripotency associated surface marker, Epithelial Cell Adhesion Molecule (EpCAM), is also expressed in the human fetal gonads and can be used as an effective selection marker for germ cell enrichment from differentiating hESCs. Combining OCT4 and EpCAM selection can further enrich the meiotic-competent germ cell-like cell population. Also, with the percentage of OCT4+/EpCAM+ cells as readout, we demonstrated the synergistic effect of BMP4/pSMAD1/5/8 and WNT3A/β-CATENIN in promoting hESCs toward the germline fate. Combining BMP4/WNT3A induction and OCT4/EpCAM selection can significantly increase the germ cell-like population with meiotic competency. Co-transplantation of these cells with dissociated mouse neonatal ovary cells into SCID mice resulted in more germ cell cluster formation in vivo. The stepwise platform established in this study provides a useful tool to elucidate the molecular mechanisms of human germ cell development, which has implications not only for human fertility research but regenerative medicine in general. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-17T00:11:02Z (GMT). No. of bitstreams: 1 ntu-101-D96642011-1.pdf: 8480052 bytes, checksum: c80466daf155e21831f372af48edcd70 (MD5) Previous issue date: 2012 | en |
| dc.description.tableofcontents | 口試委員會審定書……………………………………………………..i
誌謝…………………………………………………………………….ii 中文摘要………………………………………………………………iii 英文摘要……………………………………………………………….iv 第一章 Introduction 1.1 General introduction………………………………………...1 1.2 Germline development………………………………………...2 1.3 Germ cell differentiation from embryonic stem cells.....................................................4 1.4 Isolation and induction methods for embryonic stem cell derived germ cells...................................5 1.5 Rational and Significance……………………………......9 第二章 Heterogeneous early germ cell population isolated from human embryonic stem cells via OCT4-EGFP selection 2.1 Introduction ……………………………………………………11 2.2 Materials and Methods………………………………..………12 2.3 Results………………………………………………………………...21 2.4 Discussion…………………………………………………………....44 第三章 Meiotic competent human germ cell-like cells derived from human embryonic stem cells induced by BMP4/WNT3A signaling and OCT4/EpCAM selection 3.1 Introduction……………………………………………………………53 3.2 Materials and Methods……………………………………...55 3.3 Results…………………………………………………………………60 3.4 Discussion……………………………………………………………..78 第四章 Discussion……………………………………………………………..84 參考文獻……………………………………………………………..93 Curriculum Vitae…………………………………………………..101 附錄………………………………………………………………..106 | |
| dc.language.iso | en | |
| dc.title | 以BMP4/WNT3A誘導與OCT4/EpCAM之篩選可促進人類胚胎幹細胞分化為生殖細胞 | zh_TW |
| dc.title | BMP4/WNT3A induction and OCT4/EpCAM selection facilitate germ cell differentiation from human embryonic stem cells | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 100-2 | |
| dc.description.degree | 博士 | |
| dc.contributor.oralexamcommittee | 曾啟瑞,鄭登貴,何弘能,陳信孚,林國儀 | |
| dc.subject.keyword | 胚胎幹細胞,全能幹細胞,生殖細胞系,分化,OCT4,EpCAM, | zh_TW |
| dc.subject.keyword | Embryonic stem cells,Pluripotent stem cells,Germline,Differentiation,OCT4,EpCAM, | en |
| dc.relation.page | 134 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2012-07-13 | |
| dc.contributor.author-college | 生物資源暨農學院 | zh_TW |
| dc.contributor.author-dept | 生物科技研究所 | zh_TW |
| dc.date.embargo-lift | 2300-01-01 | - |
| Appears in Collections: | 生物科技研究所 | |
Files in This Item:
| File | Size | Format | |
|---|---|---|---|
| ntu-101-1.pdf Restricted Access | 8.28 MB | Adobe PDF |
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