2004 Fiscal Year Final Research Report Summary
Acceleration of bone regeneration by overexpression of vascular endothelial growth factor gene in bone marrow mesenchymal stem cells
| Project/Area Number |
15591573
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
Orthopaedic surgery
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| Research Institution | Nagoya University |
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Principal Investigator |
KITOH Hiroshi Nagoya University, Graduate School of Medicine, Research Associate, 大学院・医学系研究科, 助手 (40291174)
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| Co-Investigator(Kenkyū-buntansha) |
KITAKOJI Takahiko Nagoya University, University Hospital, Assistant Professor, 医学部附属病院, 講師 (10303637)
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| Project Period (FY) |
2003 – 2004
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| Keywords | bone and cartilage regeneration / gene transfer / SOX9 / marrow derived mesenchymal stem cells / cell culture / distraction osteogenesis / platelet rich plasma |
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| Research Abstract |
(1)Mouse sox9 cDNA was transfected into bone marrow derived mesenchymal stem cells(MSCs) and chondrogenic differentiation was evaluated. In vitro high density micromass culture of these sox9 transfected MSCs demonstrated that a matrix-rich aggregate was positively stained by Alcian blue and type II collagen. Next, sox9 transfected MSCs were loaded into the diffusion chamber and transplanted into athymic mice. A massive tissue formation was visible in the chamber after 4 weeks transplantation. Histological examinations demonstrated that both Alcian blue and type II collagen were positively stained in the extracellular matrix of the mass while type X collagen was not stained.
(2)Osteogenic potential of serially passaged rat MSCs was evaluated. Osteogenic differentiation in vitro was evaluated by the concentration and mRNA expression of alkaline phosphatase and osteocalcin. For in vivo osteogenesis, MSCs in various degrees of differentiation were implanted into the athymic mice. Although e
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levated levels of osteogenic markers were prominent in the less passaged MSCs continuously cultured with osteogenic supplements, they decreased with passaging. Abundant bone and cartilage formations inside the membrane were observed in the P0 through P2 cells. In the P3 cells, however, the chambers were filled with fibrous tissues.
(3)Clinical results of distraction osteogenesis with transplantation of MSCs and platelet rich plasma(PRP) were reviewed in five femora and three tibiae of the three patients with achondroplasia and two patients with leg length discrepancy. MSCs derived from the iliac crest were cultured with osteogenic supplements and differentiated into osteoblast-like cells. PRP was prepared just before transplantation. Culture expanded osteoblast-like cells and autologous PRP were injected into the distracted callus with the thrombin-calcium mixture so that the PRP gel might develop within the injected site. The target lengths were obtained in every leg without major complications and the average healing index was 24.0 day/cm. Less
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