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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 黃筱鈞(Hsiao-Chun Huang) | |
| dc.contributor.author | Shang-Fu Chen | en |
| dc.contributor.author | 陳尚甫 | zh_TW |
| dc.date.accessioned | 2021-07-11T14:39:18Z | - |
| dc.date.available | 2022-10-03 | |
| dc.date.copyright | 2017-10-03 | |
| dc.date.issued | 2017 | |
| dc.date.submitted | 2017-04-13 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/77999 | - |
| dc.description.abstract | 透過細胞再編程將人類體細胞誘導為幹細胞，使得探討細胞命運決定的方式得以獲得革新性的進步，並迎來個人化再生醫療的嶄新紀元。其中，以人類間質幹細胞於臨床細胞再生療法的應用潛力最具前瞻性。然而，成人體內的間質幹細胞不僅難以萃取、含量也相當低，成了相關自體細胞療法的一大瓶頸。為此，本團隊日前首創以複合六個小分子雞尾酒試劑，於六天內便將人體皮膚纖維母細胞化學誘導為大量間質幹細胞。此誘導式間質幹細胞更符合了所有傳統國際規範之間質幹細胞定義。該初步成果遂已於近期發表在國際期刊《科學報導》。於此，本研究首先量化誘導間質幹細胞之數個幹細胞標準能力。並藉由提升轉換效率，優化前作之雞尾酒試劑。先探討其中各單一藥物的地位，鑑定出其中的必需因子Go6983後，大量試驗120種以上的組合以尋找新試劑，結果發現四到八種藥物組成新試劑表現最佳。不但將產率由38% 提高至78%，更可省略細胞純化的步驟，而產生具骨化能力之轉分化細胞，暗示了優化試劑提高老化人體間質幹細胞分化能力的潛力。因此，本研究未來首重以優化試劑處理老化癥狀之人體間質幹細胞，測試其細胞回春之效益。總而言之，本研究可謂開拓了小分子化合物轉分化產生誘導式間質幹細胞的新篇章，指日可待其貢獻於幹細胞研究與再生醫學療法。 | zh_TW |
| dc.description.abstract | The induction of stem cells from human somatic cells through cellular reprogramming has revolutionized the way we study cell-fate decision and is heralding a new era of patient-specific regenerative medicine. Within several types of stem cells, human mesenchymal stromal/stem cells (hMSCs) hold great promises for regenerative medicine and perspectives in cell therapies. In terms of their multipotency, immuno-modularity, and without oncogenicity, hMSCs meet with high-class safety standard and have been attempted or ongoing to treat dozens of diseases through hundreds of clinical trials. However, hMSCs remain rare in adult bodies and require invasive treatment to isolate them, which is the main obstacle for autologous hMSC-based therapies. We recently demonstrated the first method to convert human dermal fibroblasts directly into functional MSC-like cells (induced MSCs, iMSCs) by a footprint-free method that avoiding DNA insertional mutagenesis. The method is based on a cocktail of six chemicals and the conversion time is within 6 days. iMSCs fulfill all the criteria of traditional hMSCs as determined by the Mesenchymal and Tissue Stem Cell Committee of the International Society for Cellular Therapy (ISCT), including cell adhesion, marker expression and multipotency (ability to differentiate into osteoblasts, adipocytes, and chondrocytes) in a degree comparable to primary human bone marrow MSCs (BMMSCs). The preliminary data proved its significance and contribution to stem cell biology and regenerative medicine, and the results were published in Scientific Reports recently. Herein, in this thesis, I firstly identified the role of single factor included in the 6C cocktail (SB202190, SP600125, Go6983, Y27632, PD0325901, and CHIR99021) and found G06983 is the essential factor for reprogramming. After testing 120 different combinations, I then boosted the conversion rate from 38% to 78% based on a concise cocktail including the essential factor. Of note, the optimized cocktails including 4C (Go6983, CHIR99021, valproic acid (VPA), and dorsomorphin), 7C (6C plus VPA), 8C cocktail (7C plus dorsomorphin) show a promising performance of improving the functions of hMSCs. Accordingly, I will treat the senescent or aging primary hMSCs with the cocktails to test if the cocktails could improve the differentiation function of aging hMSCs in the future. Overall, my findings reveal a plastic strategy to enrich functional iMSCs with high efficiency from different sources by small molecule optimization, implying the potentials for cell fate remodeling and rejuvenation research. Thus, we expand the repertoire of applying small molecules to generate iMSCs from accessible cell types and ameliorate senescent/aging hMSCs, which have major implications for broader contribution in stem cell research and cell therapy of regenerative medicine. | en |
| dc.description.provenance | Made available in DSpace on 2021-07-11T14:39:18Z (GMT). No. of bitstreams: 1 ntu-106-R04b43027-1.pdf: 4061882 bytes, checksum: fb09fcff1cc3b31211c110681982d90d (MD5) Previous issue date: 2017 | en |
| dc.description.tableofcontents | 致謝 i
中文摘要 iii Abstract iv Table of Contents vii List of Figures and Tables ix 1. Introduction 1 1-1. Overview of induced stem cell biology 1 1-1-1. Reprogramming autologous stem cells from somatic cells 1 1-1-2. Chemical induction of stem cells 2 1-2. Overview of human mesenchymal stem cells (hMSCs) 3 1-2-1. Function of hMSCs 3 1-2-2. Clinical application of hMSCs 4 1-2-3. Generation of chemical induced MSCs (iMSCs) 6 2. Materials and methods 7 2-1. Reagents. 7 2-2. Cell culture 7 2-3. Flow cytometry and fluorescence-activated cell sorting (FACS) 8 2-4. Adipogenic differentiation. 9 2-5. Oil Red O staining. 9 2-6. Chemical induction 10 2-7. CFU-F assay. 10 2-8. Osteogenic differentiation. 11 2-9. Alkaline phosphatase activity assay. 11 2-10. Alizarin Red S staining 12 2-11. Statistical analysis. 12 3. Results 14 3-1. Functional characteristics of MSCs 14 3-1-1. SSEA-4 and PODXL act as functional markers to divide the population of BMMSCs with different adipogenesis ability. 14 3-1-2. iMSCs, like BMMSCs, has high clonogenicity. 14 3-2. Optimization the 6C cocktails 15 3-2-1. Go6983 acts as the essential factor in the conversion of iMSCs from human dermal fibroblasts. 15 3-2-2. Single factor in 6C cocktail can partially improve OCT4 expression and clonogenicity 16 3-2-3. Histone deacetylase (HDAC) inhibitor significantly improves the conversion rate of iMSCs as a promoting factor 17 3-2-4. Pairing Go6983 and VPA as the concise cocktail to find Dorsomorphin as the new factors able to efficiently support the conversion 19 3-2-5. Optimized chemical cocktails reach the conversion rate up to 70%. 20 3-2-6. Induced cells present osteogenesis ability without sorting in the optimized protocol. 20 4. Discussion 22 5. Conclusion and future work 33 6. Reference 35 7. Appendix 83 | |
| dc.language.iso | en | |
| dc.title | 以藥物優化再編程人類體細胞成為誘導間質幹細胞之製程 | zh_TW |
| dc.title | Optimize the Methods of Reprogramming Chemically Induced Mesenchymal Stem Cells from Human Somatic Cells | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 105-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.coadvisor | 呂仁(Jean Lu) | |
| dc.contributor.oralexamcommittee | 林劭品(Shau-Ping Lin),陳佑宗(You-Tzung Chen),蔡素宜(Su-Yi Tsai) | |
| dc.subject.keyword | 小分子化合物,再編程,回春作用,人類間質幹細胞,再生醫學,細胞療法, | zh_TW |
| dc.subject.keyword | Small molecules,Reprogramming,Rejuvenation,Human mesenchymal stem/stromal cell,Regenerative medicine,Cell therapy, | en |
| dc.relation.page | 84 | |
| dc.identifier.doi | 10.6342/NTU201700753 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2017-04-14 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 分子與細胞生物學研究所 | zh_TW |
| 顯示於系所單位： | 分子與細胞生物學研究所 | |
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