2004 Fiscal Year Final Research Report Summary
Ex Vivo Gene Therapy for Dentin Regeneration Using Dental Pulp Stem Cells transfected with Gdf11
| Project/Area Number |
15390577
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Conservative dentistry
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| Research Institution | KYUSHU UNIVERSITY |
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Principal Investigator |
NAKASHIMA Misako Kyushu University, Faculty of Dental Science, Research Associate, 大学院・歯学研究院, 助手 (20207773)
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| Co-Investigator(Kenkyū-buntansha) |
AKAMINE Akifumi Kyushu University, Faculty of Dental Science, Professor, 大学院・歯学研究院, 教授 (00117053)
HIRAYAMA Yoshinori The University of Kitakyushu, Faculty of Environmental Engineering, Professor, 国祭環境工学部, 教授 (40253504)
HIRATA Masako Kyushu University, Hospital, Research Associate, 大学病院, 助手 (10153769)
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| Project Period (FY) |
2003 – 2004
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| Keywords | Gene Therapy / Dentin / Pulp Regeneration / Reparative Detin / Dental Pulp Stem Cells / Side Population Cells / Odontoblasts / Bone Morphogenetic Proteins (BMPs) / Flow Cytometry |
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| Research Abstract |
The regeneration of dentin is the goal of operative dentistry. The key elements of regenerative medicine are stem cells, morphogens and scaffold of extracellular matrix such as collagen. The dental pulp has progenitor/stem cells that have the potential to differentiate into dentin-forming odontoblasts in response to members of bone morphogenetic proteins (BMPs), morphogen family. BMPs are inductive morphogenetic signals. In this investigation, we have developed a successful ex vivo gene therapy for regenerative dentin formation using pulp stem cells and a morphogen of the BMP family, Gdf11/Bmp11. Side population (SP) cells, isolated from adult pulp tissue based on the exclusion of the DNA dye Hoechst 33342, exhibited stem cell activity, such as self-renewal capability and multipotency to differentiate into neurons, chondrocytes and adipocytes as well as odontoblasts. The expression of Bmil, Stat3 and Tert mRNA, the markers for stem cells is increased in the SP cells compared with non-SP cells. The pulp stem cells were electrotransfected with Gdf11. The transfection efficiency was nearly 70%. Real time RT PCR analysis confirmed the elevated expression of odontoblast-specific markers such as ALPase, dentin matrix protein 1 (Dmp1), dentin sialophosphoprotein (Dspp), enamelysin and phosphate-regulating gene with homologies to endopeptidases on X-chromosome (Phex) by the Gdf11 transfected cells. Based on this promising in vitro proof-of-concept experiment, an in vivo evaluation in the dog was undertaken using autogenous canine pulp cells transfected with Gdf11. Transplantation of Gdf11-transfected pulp cells as a pellet on surgically amputated pulp induced regenerative dentin formation as evaluated by radiographs and histology. In conclusion, the transfection of dentin morphogen, Gdf11 into pulp progenitor/stem cells can be used for regenerative therapy of dentin for endodontic therapy in dentistry.
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