Optimization of ectopic bone marrow formation following transplantation of mesenchymal stem cell expressing bone morphogenic factor-2 and endothelial cells

다국어 초록 (Multilingual Abstract)

Bone marrow (BM) is an important immune organ which contains hematopoietic cells and consists of adventitial reticular cells, osteocytes, adipocytes, extracellular collagen matrix, and vascular endothelium. BM microenvironment plays a critical role in hematopoiesis. BM niche is important for human hematopoietic stem cell (HSC) engraftment.
Over the past few decades, mesenchyal stem cells (MSCs) have become of increasing for use in the field of regenerative medicine. Several groups used MSCs for making BM microenvironment, because MSCs have ability to differentiate into osteocytes, osteoblasts, chondrocytes, endothelial cells and so on. Also, bone morphogenetic protein 2 (BMP2), a member of large family of proteins, has been known for its ability to induce the formation of bone and cartilage.
We describe the development of ectopic bone marrow using human autologous BMP2 expressed umbilical cord (UC) derived MSCs and umbilical cord derived endothelial cells (ECs) implanted subcutaneously into immunodeficient mice. We could establish human ectopic bone marrow construct in the NOD-scid IL2rγnull (NSG) subcutaneous. This model could be useful for the study of the development of novel therapeutic modalities aimed at modifying the hematopoietic microenvironment. Also, Autologous BM oganogenesis will become an invaluable tool for human HSC engraftment.

목차 (Table of Contents)

• 1. ABSTRACT 1
• 2. INTRODUCTION 3
• 3. MATERIALS AND METHODS 6
• 3-1. Isolation of the umbilical cord derived MSCs and the culture 6
• 3-2. Immunophenotyping of the umbilical cord derived MSCs by flow cytometry 6
• 3-3. Osteogenic differentiation 7
• 3-4. Adipogenic differentiation 7
• 3-5. Isolation of the umbilical cord derived ECs and the culture 8
• 3-6. Immunophenotyping of the umbilical cord derived ECs by flow cytometry 8
• 3-7. Umbilical cord derived ECs tube formation functional assay 9
• 3-8. In vitro adenovirus transduction 9
• 3-9. BMP2 enzyme-linked immunosorbent assay (ELISA) analysis 10
• 3-10. Western blot analysis 10
• 3-11. Apoptosis analysis 10
• 3-12. Immunohistochemistry, immunofluorescence analysis 11
• 4. RESULTS 12
• 4-1. Umbilical cord derived MSCs characterization and differentiation assay 12
• 4-2. Umbilical cord derived ECs characterization and functional assay 12
• 4-3. Optimization of soluble BMP2 production in Umbilical cord derived MSCs 13
• 4-4. Ectopic bone marrow analysis 14
• 5. DISCUSSION 24
• 6. CONCLUSION 26
• 7. REFERENCES 27
• 8. 국문초록 31