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인간 귀 유양골수 및 지방조직 유래 줄기세포를 이용한 유양골 분화 유도

초록/요약

Injured bones have a limited capacity for self-repair in most cases, including otolaryngology. In the case of patients with chronic otitis media or people who need a cochlear implant surgery, the mastoid bone must be drilled out. However, it results in temporal bone defects which can cause various complications such as dizziness, accumulation of keratin debris, otorrhea, or recurrent lesions. To compensate for these disadvantages of open cavity mastoidectomy, reconstruction of the posterior wall of the external auditory canal and mastoid obliteration have been attempted. Autologous tissues, alloplastic and biosynthetic materials have all been used in mastoid obliteration. Autologous tissues are much safer and more cost-effective than alloplastic or biosynthetic materials. However, they also have limitations and are not applicable in every situation because of insufficient amounts of autologous tissues, donor site defects, and reabsorption after surgery. Mastoid bone marrow and adipose tissue can be easily obtained during the surgery without any additional invasive procedures, and also be great source of stem cell. Mastoid bone is normally characterized with air cells for gas exchange and also remains empty space after mastoidectomy. Therefore, mastoid bone requires different bone regeneration strategy from compact bones including cranial bones. Naturally, the scaffold may have to be different depending on the kinds of target bones. Artificial scaffolds must also be biodegradable, allowing extracellular matrix to occupy the empty space when the biomaterial is degraded. The aim of this study was to investigate the effectiveness of mastoid bone marrow and adipose tissue-derived stem cells, which are acquired through mastoidectomy, and to evaluate their effects on bone formation in combination with scaffold to regenerate temporal bone defects. The human mastoid bone marrow and adipose tissues showed stem cell phenotypes, and these characteristics were maintained up to passage 5. Mastoid bulla and cranial bone defects were induced in Sprague-Dawley rats and the rats were divided into five groups: (1) control, (2) human ear adipose-derived stromal cells (hEASCs), (3) hEASCs + osteogenic differentiation medium (ODM), (4) hEASCs + polycaprolactone (PCL) scaffolds, and (5) hEASCs + PCL scaffolds + ODM. Osteogenesis was evaluated by micro-computed tomography and histology. Compared with the control group, the groups transplanted with hEASCs and PCL scaffolds had significantly higher bone formation along the periphery of the mastoid bulla area. Moreover, ODM synergistically enhanced bone formation in mastoid bulla defects. Our results suggest that combining hEASCs with PCL scaffolds represents a promising method for anatomical and functional reconstruction of postoperative temporal bone defects following mastoidectomy

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목차

Ⅰ. INTRODUCTION 1
Ⅱ. MATERIALS AND METHODS 5
1. Cell isolation and culture 5
2. Fluorescent activated cell sorting analysis 8
3. Multi-lineage differentiation of cells 8
4. Cytochemistry 9
5. Immunofluorescence 9
6. Western blot 10
7. Histological analysis 10
8. Animal experimental design 11
9. PCL 3D scaffold 12
10. Surgery 14
11. Micro-CT 14
12. Statistical analysis 15

Ⅲ. RESULTS 16
1. Clinical profiles of the patients 16
2. Characterization of mastoid bone marrow-derived cells 18
3. Maintenance of stem cell phenotype in mastoid bone marrow-derived cells 20
4. Differentiation of mastoid-derived cells into the chondrogenic, osteogenic, adipogenic, neurogenic and epithelial Lineages 22
5. Characterization of hEASCs 26
6. Bone formation in rat mastoid bulla and cranial bone defects 28
7. ODM synergistically enhances cell/scaffold-based bone formation 31
Ⅳ. DISCUSSION 34
Ⅴ. CONCLUSION 39
REFERENCES 40
국문요약 45

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