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http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/78878完整後設資料紀錄
| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 黃義侑 | |
| dc.contributor.author | Hung-Jun Lin | en |
| dc.contributor.author | 林宏潤 | zh_TW |
| dc.date.accessioned | 2021-07-11T15:26:06Z | - |
| dc.date.available | 2023-11-08 | |
| dc.date.copyright | 2018-11-08 | |
| dc.date.issued | 2018 | |
| dc.date.submitted | 2018-11-06 | |
| dc.identifier.citation | [1] C.C. Teng, Fine structure of the human cornea: epithelium and stroma, Am J Ophthalmol 54 (1962) 969-1002.
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/78878 | - |
| dc.description.abstract | 近年來全球人口老化及罹患疾病趨勢提高，以再生醫學重建人體健康是熱門的研發策略，而組織工程結合幹細胞治療被視為最具臨床應用潛力的重要研究。去細胞支架具有天然的組織結構及生物因子，適合在體外建立最佳的培育條件，因此值得深入研究其臨床應用方式。本研究建立有機酸去細胞組織工程技術，針對角膜再生及樹突狀細胞支架癌症免疫療法之應用潛力進行相關探討。
將豬眼角膜以有機酸去細胞方式得到去細胞眼角膜支架，結果顯示甲酸能清除支架內多數細胞，並保留支架內膠原蛋白和葡萄糖胺聚合醣。此外，去細胞眼角膜支架呈現與原有結構相似之機械強度及平行薄板結構。在細胞實驗顯示，去細胞眼角膜支架具有良好生物相容性，角膜上皮細胞能適當的生長於支架上，而甲酸去細胞創造之支架孔洞可利於角膜基質細胞存活於去細胞眼角膜支架內。動物眼角膜移植試驗證實，甲酸去細胞眼角膜支架植入動物後，角膜上皮細胞能快速生長佈滿其支架表面，角膜基質細胞能進入支架內生長並釋放基質填滿內部孔洞，且能長時間維持角膜原有透明度。因此本研究證實，以有機酸去細胞技術建立之去細胞眼角膜支架，具有應用於臨床角膜移植潛力。 在癌症免疫治療應用方面，本研究建立帶有活化樹突狀細胞之去細胞淋巴結支架，植入體內後觀察其抑制癌細胞生長之能力。結果顯示甲酸對於大鼠淋巴結能有效去除原有細胞，對於基質內膠原蛋白及葡萄糖胺聚合醣能有效保留。而樹突狀細胞可存活於去細胞淋巴結支架內，經抗原OVA (Ovalbumin)刺激後活化成熟標記CD80、CD86和MHC-Ⅱ分子，並釋放細胞激素IL-1β、IL-6和IL-12。此外，將帶有樹突狀細胞之去細胞淋巴結支架與CD8+毒殺型T淋巴球共同培養後，對於細胞激素IL-1β、IL-6、IL-12和IFN-γ能顯著提升其釋放量，這有助於後續免疫反應強度的提升。動物腫瘤實驗證實，給予抗原活化樹突狀細胞之去細胞淋巴結支架免疫後之小鼠，能完全抑制E.G7腫瘤細胞的生長，具有良好的免疫預防效果。而在取出免疫後小鼠脾臟細胞經由體外給予抗原OVA再刺激後，細胞激素IFN-γ、IL-4、IL-6和IL-2有明顯提升，且脾臟細胞增生率也有增加的情形。這些結果證實，抗原活化樹突狀細胞之去細胞淋巴結支架可誘發抗原專一性免疫反應，對於動物之腫瘤細胞能有效抑制其生長，因此在臨床癌症細胞免疫治療具有應用價值。 | zh_TW |
| dc.description.abstract | In recent decades, decellularized extracellular matrix (ECM) showed its potential as a promising scaffold used in the fields of tissue regeneration. In this study, organic acid decellularized technique was evaluated for corneal regeneration and DC (dendritic cells)-based scaffold cancer vaccine.
For corneal regeneration, the decellularized porcine corneal scaffolds (dPC) were prepared by organic acid treatment. Cell removal and intact extracellular matrix preservation was evidenced by histological and biochemical quantitative analysis, and the dPC scaffolds showed porous parallel lamellar microstructure and great biomechanical properties. In-vitro cell culture demonstrated that the dPC scaffolds had good biocompatibility, and the porous microstructure provided an ideal space for the growth of stroma keratocytes. Moreover, in-vivo implantation revealed ideal re-epithelialization, stromal recellularization and complete transparency during the full follow-up period. Thus, dPC scaffolds that prepared by organic acid treatment could be a promising biological material for use in corneal transplantation. For cancer immunotherapy, the decellularized lymph node scaffolds (dLN) also prepared by organic acid treatment. The results showed the highly efficient removal of cell debris from lymph node, and great preservation of ECM architecture and biomolecules. In addition, bone marrow dendritic cells (BMDC) grown preferably inside the dLN, displayed maturation marker CD80, CD86 and MHC-Ⅱ, and produced high level of cytokine IL-1β, IL-6 and IL-12 when stimulated with ovalbumin (OVA) and CpG oligodeoxynucleotides (CPG-ODN). Furthermore, BMDC-dLN co-cultured with CD8+ T lymphocytes revealed higher production of IL-1β, IL-6, IL-12 and IFN-γ. In animal model, the BMDC-dLN showed completely rejected the E.G7-OVA tumor, and the splenocytes from BMDC-dLN immunized mice produced more IFN-γ, IL-4, IL-6 and IL-2, and higher proliferation rate than other groups when re-stimulated with OVA. Hence, this could be a promising DC-based scaffold for delivery in-vivo to induce potent anti-tumor immunity. | en |
| dc.description.provenance | Made available in DSpace on 2021-07-11T15:26:06Z (GMT). No. of bitstreams: 1 ntu-107-D02548004-1.pdf: 3810461 bytes, checksum: c075d311add7b45fab0e043044c40568 (MD5) Previous issue date: 2018 | en |
| dc.description.tableofcontents | List of Contents
論文口試委員審定書 i 誌謝 ii 中文摘要 iv Abstract vi List of Contents viii List of Figures xiii Abbreviations xiv Introduction 1 Part 1 Development of Decellularized Cornea for Corneal Regeneration 2 Chapter 1 Background 3 1.1 Cornea 3 1.1.1 Anatomy and functions of cornea 3 1.1.2 Cornea transplantation 4 1.2 Scaffold types used in corneal tissue engineering 5 1.2.1 Synthetic corneal scaffolds 6 1.2.2 Biological corneal scaffolds 7 1.2.3 Decellularized Corneal Scaffolds 7 1.3 The Aim of this research 9 Chapter 2 Materials and Methods 10 2.1 Materials 10 2.2 Decellularization of porcine corneas 11 2.3 Assessment of the optical property of dPC scaffolds 12 2.4 Mechanical properties and microstructure of dPC scaffolds 12 2.5 Biochemical analysis and histological examination 13 2.6 Recellularization on dPC scaffolds 14 2.7 dPC scaffolds for corneal xenotransplantation 16 2.7 Statistical analysis 17 Chapter 3 Results 18 3.1 Transparency, mechanical properties, and microstructure of the dPC scaffolds 18 3.2 The efficacy of the decellularization process 19 3.3 Characterization of ECM components 20 3.4 In-vitro dPC scaffolds recellularization 21 3.5 Implantation of the dPC scaffolds in-vivo 22 Chapter 4 Discussion 31 4.1. The comparison of decellularized methods 31 4.2. Re-cellularization of corneal epithelial cells and keratocytes on dPC 32 4.3. Corneal regeneration of dPC on rabbit model 33 Chapter 5 Conclusions 35 Part 2 Development of Decellularized Lymph node for Cancer immunotherapy 36 Chapter 6 Background 37 6.1 Lymph node 37 6.1.1 Anatomy and functions of lymph node 37 6.1.2 Lymphedema 39 6.1.3 Lymph node transfer 40 6.2 Immune system 41 6.2.1 Adaptive immune system 42 6.2.2 Dendritic cells 43 6.2.3 T lymphocytes 44 6.3 Cancer immunotherapy 45 6.4 Vaccine adjuvant 47 6.5 The aim of this research 48 Chapter 7 Materials and Methods 49 7.1 Materials 49 7.2 Decellularization of lymph node 50 7.3 Microstructure of dLN scaffolds 50 7.4 DNA quantification analysis and histological examination 51 7.5 Recellularization of BMDC into dLN scaffolds 52 7.6 Stimulation of BMDC-dLN and the cytokine profile 53 7.7 BMDC co-cultured with CD8+ T lymphocytes in dLN 53 7.8 Immunization of mice and tumor challenge 54 7.9 Analysis of cells population inside the BMDC-dLN after immunization 55 7.10 Ex-vivo re-stimulation of splenocytes 55 7.11 Statistical analysis 56 Chapter 8 Results 57 8.1 The efficacy of the decellularization process 57 8.2 Microstructure and ECM examination of the dLN scaffolds 57 8.3 Maturation marker analysis and cytokine released of BMDC-dLN 58 8.4 CD8+ T lymphocytes co-cultured with BMDC-dLN 60 8.5 Protective effect of BMDC-dLN vaccine against tumor challenge 61 8.6 Analysis of cell population in BMDC-dLN after immunization 61 8.7 Ex-vivo re-stimulation of splenocytes 62 Chapter 9 Discussion 78 9.1 The dLN scaffolds provided suitable 3D environment 78 9.2 Cytokine production ability of the BMDC-dLN 80 9.3 In-vivo anti-tumor effects of the BMDC-dLN 82 Chapter 10 Conclusions 84 References 85 | |
| dc.language.iso | en | |
| dc.title | 去細胞組織工程應用於眼角膜再生醫學及癌症免疫治療之研究 | zh_TW |
| dc.title | Application of Decelluarized Tissue Engineering for Corneal Regeneration and Cancer Immunotherapy | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 107-1 | |
| dc.description.degree | 博士 | |
| dc.contributor.oralexamcommittee | 劉得任,鍾次文,許明照,張育嘉,王宗仁 | |
| dc.subject.keyword | 組織工程,去細胞支架,角膜再生,癌症免疫治療,樹突狀細胞, | zh_TW |
| dc.subject.keyword | Tissue engineering,decellularized scaffold,corneal regeneration,cancer immunotherapy,dendritic cell, | en |
| dc.relation.page | 94 | |
| dc.identifier.doi | 10.6342/NTU201804231 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2018-11-07 | |
| dc.contributor.author-college | 工學院 | zh_TW |
| dc.contributor.author-dept | 醫學工程學研究所 | zh_TW |
| dc.date.embargo-lift | 2023-11-08 | - |
| 顯示於系所單位： | 醫學工程學研究所 | |
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