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研究生: 陳鼎元
論文名稱: 三維細胞球體之開發於缺血性組織血管新生之應用
Development of Three-Dimensional Cell Aggregates for Therapeutic Neovascularization in Ischemic Tissues
指導教授: 宋信文
口試委員: 陳三元
王麗芳
黃效民
張燕
林昆儒
學位類別: 博士
Doctor
系所名稱: 工學院 - 化學工程學系
Department of Chemical Engineering
論文出版年: 2013
畢業學年度: 101
語文別: 英文
論文頁數: 78
中文關鍵詞: 細胞球體血管新生缺血性組織
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  • 細胞治療在組織工程與再生醫學的領域上,為一相當具有前瞻性的治療方法。我們先前與清華大學化工系宋信文教授的團隊合作,利用細胞不會貼附在甲基纖維素水膠的特性,開發出一能生產大小均一且可經針頭注射的3-D臍帶血間葉幹細胞/人類臍帶靜脈內皮細胞 (cbMSC/HUVEC) 均勻混合細胞球體生產系統。探討此均勻混合細胞球體的增生、分佈及最佳化等特性。體外實驗結果顯示,此cbMSC與HUVEC均勻混合於細胞球體內部並具有完整的細胞外間質及黏附性蛋白,可以形成穩定的類血管網狀結構,也找出促進此類血管網狀結構形成的相關機制。證實此均勻混合細胞球體確實有類血管形成的能力。在體內實驗,我們將此cbMSC/HUVEC均勻混合細胞球體利用針頭植入缺血肌肉組織進行血管新生的治療。實驗結果顯示此均勻混合細胞球體的確可以促進缺血性肌肉組織血管新生。


    ABSTRACT I TABLE OF CONTENT III LIST OF FIGURES AND TABLES VI Chapter 1. Introduction 1 Chapter 2. Study I 6 2.1. Materials and Methods 6 2.1.1. Cell culture 6 2.1.2. Cultivation and characterization of cell aggregates 7 2.1.3. Tube formation assay 8 2.1.4. Real-time polymerase chain reaction (PCR) 9 2.1.5. Animal study 9 2.1.6. Animal perfusion imaging 10 2.1.7. Histological analyses 10 2.1.8. Statistical analysis-------------------------------------------------- 11 2.2. Results and Discussion 12 2.2.1. Cultivation and characterization of cell aggregates 12 2.2.2. Tube formation assay 14 2.2.3. Colocalization of cbMSCs with HUVECs 17 2.2.4. Gene expression 18 2.2.5. Hindlimb salvage and blood perfusion 20 2.2.6. Histological analyses 23 2.3. Conclusions 26 Chapter 3. Study II 27 3.1. Materials and Methods 27 3.1.1. Preparation and characterization of cell aggregates 27 3.1.2. Tube formation assay 28 3.1.3. Animal study 29 3.1.4. SPECT imaging 29 3.1.5. Echocardiography 30 3.1.6. Morphometric and histological analyses 30 3.1.7. Statistical analysis 31 3.2. Results and Discussion 32 3.2.1. Characteristics of cell aggregates 32 3.2.2. Tube formation assay 33 3.2.3. Assessment of myocardial perfusion by SPECT 37 3.2.4. Evaluation of LV function by echocardiography 39 3.2.5. Histological analyses 42 3.3. Conclusions 45 Chapter 4. Study III 46 4.1. Materials and Methods 46 4.1.1. Cell culture 46 4.1.2. Fabrication and characterization of cell aggregates 47 4.1.3. Tube formation assay 49 4.1.4. Animal study 49 4.1.5. SPECT perfusion imaging 49 4.1.6. Histological analyses 50 4.1.7. Statistical analysis 50 4.2. Results and Discussion 52 4.2.1. Fabrication of hypoxic HUVEC/cbMSC aggregates 52 4.2.2. Characteristics of hypoxic HUVEC/cbMSC aggregates 53 4.2.3. Angiogenic potential of hypoxic aggregates 55 4.2.4. Immunostaining of hypoxic aggregates 56 4.2.5. Tube formation assay and gene expression 57 4.2.6. Recovery of muscle atrophy and blood perfusion 61 4.2.7. Histological analyses 63 4.3. Conclusions 66 References 67 Publication 78

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