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http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/31895完整後設資料紀錄
| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 侯連團(Lein-Tuan Hou) | |
| dc.contributor.author | "CHIN-LUNG, LIU" | en |
| dc.contributor.author | 劉錦龍 | zh_TW |
| dc.date.accessioned | 2021-06-13T03:23:39Z | - |
| dc.date.available | 2015-01-01 | |
| dc.date.copyright | 2006-08-04 | |
| dc.date.issued | 2006 | |
| dc.date.submitted | 2006-07-29 | |
| dc.identifier.citation | Abdennagy B, Hott M, Marie PJ (1992). Effects of platelet-derived growth factor on human and mouse osteoblastic cells isolated from the trabecular bone surface. Cell Biol Int Rep 16(3):235-47.
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/31895 | - |
| dc.description.abstract | Background: Tissue engineering for bone regeneration relies on the presence of proper growth factors inside a suitable delivery system. Bone morphogenetic proteins (BMPs) are well-known important growth factors capable of inducing ectopic bone formation. However, an effective carrier/delivery system is critical for BMPs to accomplish this result. The objective of this study was to test the in vitro and ex vivo potential of the novel composite scaffold (synthetic polymers/gelatin) as a carrier for rhBMP-2.
Materials and methods: We prepared several scaffolds for carring rhBMP-2. The synthetic polymers of poly(DL-lactide) was fabricated with gelatin to be a test composite scaffold for carrying rhBMP-2. The gelatin sponge and PDLLA scaffold (PDLLA) were also prepared for carring rhBMP-2 as controls. The structure, pore sizes, and cell affinity of test and control scaffolds were studied by scanning electron micrograph (SEM). The compression strength between test and some control scaffold was compared. Compositional change between PDLLA and gelatin in composite scaffold were evaluated Fourier Transform Infrared Spectroscopy (FTIR) analysis. In ex vivo assay, Composite, PDLLA, and gelatin loaded with 30μg rhBMP-2 ( rhBMP-2-Composite; rhBMP-2-PDLLA; rh-BMP-2-gelatin) and control scaffolds were implanted into calf muscle of 24 male Wistar rats (4-5 weeks-old) to explore the ectopic bone formation at 3, 7, 14, and 28 days. Tissue blocks were harvested and ectopic bone formation was examined by histology, Masson’s trichrome staining, Goldner’s staining, and immunohistochemistry of key bone matrix proteins (OPN and BSP). RT-PCR analysis was performed for the expression of osteoblastic gene such as core binding factor 1 (Cbfa1), alkaline phosphatase (ALP), osteopontin (OPN), and osteocalcin (OCN). Results: Under SEM observation, PDLLA scaffolds showed a dense non-porous outer surface skin and a mean pore size ＜ 10μm and poor pores, interconnection cross-sectionally.. The composite scaffolds (test) exhibited porous outer surface and a porosity (pores size 50-60μm) inside the scaffold than those of control PDLLA scaffolds. Cell affinity of the test scaffolds were also better than that of PDLLA. Composite scaffolds showed better compression strength than galatin one. FTIR analysis showed that compositional profile of the test scaffold is a mixture of PDLLA and gelatin. In ex vivo study, all implants showed good biocompatibility without severe inflammatory responses and foreign body reaction. The PDLLA-rhBMP-2 and control PDLLA scaffolds failed to induce bone formation throughout the study. Initial events of ectopic bone formation were found in Gelatin-rhBMP-2 and Composite-rhBMP-2- at 7 and 14 days, respectively. The results of RT-PCR analysis indicated that consistent gene expression of Cbfa1, ALP and OPN are found in tissue specimens implanted harvested from Composit-rhBMP-2 and Gelatin- rhBMP-2. Conclusion: Novel composite scaffolds displayed a structure with proper micro-pore structure, good tissue biocompatibility and mechanical strength. The incorporation of rhBMP-2 into composite scaffolds could induce ectopic bone formation as shown by histology, immunohistochemistry and gene expression within 2 weeks. The results suggest that the novel composite scaffolds can be a promising scaffold for carrying rhBMP-2 in tissue engineering of bone. | en |
| dc.description.provenance | Made available in DSpace on 2021-06-13T03:23:39Z (GMT). No. of bitstreams: 1 ntu-95-R92422014-1.pdf: 3551870 bytes, checksum: 84ed6baf07f4a63b08d43c372b629835 (MD5) Previous issue date: 2006 | en |
| dc.description.tableofcontents | Table of Content
ABSTRACT 8 I. REVIEW OF LITERATURE 11 A. Periodontal bioengineering 11 A.1. Current Periodontal Therapies 11 A.2. Tissue engineering approach in periodontal bone regeneration 17 i. Cells for periodontal bone tissue engineering 18 ii. Growth factors for periodontal bone tissue engineering 22 iii. Scaffolds for periodontal bone tissue engineering 29 B. Bone Morphogenetic Proteins 41 B.1. Introduction 41 B.2. General Roles of BMPs 43 B.3. Roles of BMPs on formation of the embryonic skeleton and osteogenesis in the adult skeleton 46 i. Roles of BMPs on formation of the embryonic skeleton 46 ii. Osteogenesis in adult skeleton 47 B.4. Functional mechanism and signal transduction of BMPs 49 B.5. Clinical applications of rhBMPs in dentistry 54 C. Synthetic Polymeric Biomaterials for bone tissue engineering 56 C.1. Introduction 56 C.2. Degradation of polymeric biomaterials 59 C.3. Current scaffold design and fabrication 62 C.4. Incorporation of active biomolecule(s) onto polymeric biometerials 68 II. HYPOTHESIS AND SPECIFIC AIM OF THIS STUDY 70 III. INTRODUCTION 72 IV. MATERIALS AND METHODS 77 A. In vitro study 77 A.1. Preparation of PDLLA and composite scaffolds with or without rhBMP-2 77 A.2. Structural and cell affinity analysis of scaffolds under scanning electron micrograph (SEM) 79 A.3. Fourier Transform Infrared Spectroscopy (FTIR) analysis 80 A.4. Compression strength analysis 80 B. Ex vivo study 82 B.1. Animal study 82 i. Experiment I: The PDLLA scaffold impregnated with rhBMP-2 82 ii. Experiment II: The Composite scaffold impregnated with rhBMP-2 83 B.2. Histological examination 83 i. Hematoxylin and Eosin stain 84 ii. Masson’s Trichrome stain 84 iii. Goldner’s Stain 84 B.3. Immunohistochemical staining 84 B.4. Semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR) detection for genes expression of bone specific proteins of tissue biopsies. 85 V. RESULTS 88 A. In vitro study 88 A.1. Structure and cell affinity analysis of scaffolds under SEM 88 A.2. FTIR analysis 89 A.3. Compression strength test analysis 90 B. Ex vivo study 91 B.1. Gross and stereoscope examination 91 B.2. Histological examination and Immunohistochemical staining 91 i. Experiment I: The PDLLA scaffold impregnated with rhBMP-2 91 a) H&E staining 91 b) Masson’s trichrome staining 93 c) Goldner’s staining 94 d) Immunohistochemical staining, OPN and BSP 94 ii. Experiment II: The Composite scaffold impregnated with rhBMP-2 95 a) H&E staining 95 b) Masson’s trichrome staining 99 c) Goldner’s staining 100 d) Immunohistochemical staining, OPN and BSP 102 B.3. RT-PCR detection for genes expression of bone specific proteins of tissue biopsies 104 VI. DISCUSSION 105 A. Scaffold characteristics and bone tissue engineering 105 A.1. Biocompatibility and degradation of the scaffold 105 A.2. Pore size of the scaffold 106 A.3. The scaffolds’ surface properties and cell affinity 108 B. Gene expression of bone specific marker, and bone formation induced by BMPs 110 C. Source of cells incorporated in BMPs-induced ectopic bone formation 115 VII. CONCLUSION 120 VIII. APPENDIX 122 1. H&E stain protocol 122 2. Masson’s Trichrome stain protocol 122 3. Goldner’s stain protocol 124 4. Immunohistochemical staining protocol 126 IX. REFERENCE LIST 174 List of Figures Review of literature: Fig 1: Crosstalk of the BMP–Smad pathway with the TGF-b/activin signal transduction pathways 51 Fig 2: Signal transduction of BMPs through Smad pathway 52 Fig 3: BMP-smad and BMP-MAPK pathway 53 Fig 4: Nonspecific hydrolytic cleavage of ester bonds begins upon contact with water 61 Fig 5. Routes elimination for commonly used polyhydroxyacids 61 Results: Fig1-4: Structure and cell affinity analysis of scaffolds under SEM 129 Fig.5: FTIR analysis of PDLLA, gelatin and composite scaffold 131 Fig.6: Compression strength test analysis of composite scaffold and gelatin carrier 132 Fig.7-197: Histological examination and immunohistochemical staining 133 Fig.198: RT-PCR detection for genes expression of bone specific proteins of tissue biopsies 165 LIST OF TABLES Review of literature Table 1. BMP family in human and chromosome location 42 Results: Table 1. Primers of bone-related key proteins and RT-PCR condition 166 Table 2. Summery of histological examination in PDLLA scaffold 167 Table 3. Summery of histological examination in PDLLA-rhBMP-2 scaffold 168 Table 4. Summery of histological examination in composite scaffold 169 Table 5. Summery of histological examination in Composite-rhBMP-2 scaffold 170 Table 6. Summery of histological examination in Gelatin-rhBMP-2 scaffold 171 Table 7. Summery of histological examination of osteogenesis in experiment I 172 Table 8. Summery of histological examination of osteogenesis in experiment II 173 | |
| dc.language.iso | en | |
| dc.title | 合成聚合物-明膠複合體作為骨成形蛋白載體於骨再生研究中之評估 | zh_TW |
| dc.title | Evalution of Synthetic Polymers-Gelatin Composite Scaffold As rhBMP-2 Delivery System in Bone Regeneration | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 94-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.coadvisor | 劉步遠(Bu-Yuan Liu) | |
| dc.contributor.oralexamcommittee | 傅鍔(Earl Fu),王大銘(Da-Ming Wang) | |
| dc.subject.keyword | 組織工程,載體,骨成形蛋白,骨再生,合成聚合物,複合載體, | zh_TW |
| dc.subject.keyword | Tissue engineering,bone morphogenetic protein,Atrisorb,composite scaffold,PDLLA,bone induction,endochondral bone formation, | en |
| dc.relation.page | 198 | |
| dc.rights.note | 有償授權 | |
| dc.date.accepted | 2006-07-29 | |
| dc.contributor.author-college | 醫學院 | zh_TW |
| dc.contributor.author-dept | 臨床牙醫學研究所 | zh_TW |
| 顯示於系所單位： | 臨床牙醫學研究所 | |
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