Development of selective culture and cell plantation systems for mouse osteoblasts over-expressing human osteoblast stimulating factor-1 (OSF1)

开发过表达人成骨细胞刺激因子-1 (OSF1) 的小鼠成骨细胞的选择性培养和细胞种植系统

• 批准号: 14571393
• 负责人: HASHIMOTO-GOTOH Tamotsu
• 金额: $ 2.24万
• 依托单位: Kyoto Prefectural University of Medicine
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2002
• 资助国家: 日本
• 起止时间: 2002 至 2003
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-14571393/
• 关键词: osteoporosis; gene therapy; stem cell; cell plantation; 骨芽細胞

In order to establish the new therapeutic method for osteoporosis, we are investigating the model systems of cell plantation and gene therapy with osteoblast stimulating factor-1 (OSF1).The mesenchymal cells isolated from the marrow cell of the patient himself, may have and give the higher osteoblast differentiation potency when OSF1 is introduced exogenously.In order to establish model experimental system, we constructed transgenic mice expressing human OSF1 gene controlled by CAG promoter (strong ubiquitous expression) or Osteocalcine promoter (osteoblast specific expression) for the source of mesenchymal stem cells which may have high potency for osteoblast differentiation. We examined about the potency of differentiation into osteoblasts and activity of bone formation on isolated mesenchymal cells and pre-osteoblastic cells from several Tg mouse lines. We found the fact that the primary cultured osteoblasts in the presence of BMP2 exhibit the higher Ca deposition activity compared to those from non-Tg control mice. The elevation of Ca deposition activity and differentiation into osteoblasts indicated by the elevation of alkaline phosphatase activity were also observed in the case of mesenchymal cells derived from OSF1 expressing Tg-mice. Now, we are planning to evaluate the bone mass increasment effect by the transplanted cells into osteoporosis model mice.

为了建立骨质疏松症治疗的新方法，我们研究了成骨细胞刺激因子-1（osteoblasticstimulatingfactor-1，OSF-1）的细胞种植园和基因治疗的模型体系，从患者自身骨髓细胞中分离的间充质细胞，在外源性导入OSF-1后，可能具有更高的成骨分化潜能，为了建立模型实验体系，我们构建了表达由CAG启动子（强普遍表达）或Osteocalcine启动子（成骨细胞特异性表达）控制的人OSF 1基因的转基因小鼠，用于可能具有高潜能的成骨细胞分化的间充质干细胞的来源。我们研究了从几个Tg小鼠品系分离的间充质细胞和前成骨细胞分化成骨细胞的潜能和骨形成活性。我们发现，与非Tg对照小鼠相比，BMP 2存在下的原代培养成骨细胞表现出更高的钙沉积活性。在来自表达OSF 1的Tg-小鼠的间充质细胞的情况下，也观察到Ca沉积活性的升高和由碱性磷酸酶活性的升高指示的向成骨细胞的分化。目前，我们正在计划评估将细胞移植到骨质疏松模型小鼠中的骨量增加效果。

项目成果

Oshima Y, Takai S, Watanabe N, Matsuda K, Kawata M, Kubo T: "Behavior of Transplanted Bone Marrow-Derived Mesenchymal Cell-Masses during the Regenerating Process of Osteochondral Defect using Transgenic Rats"Transaction of Orthopaedic Research Society. 28

Oshima Y、Takai S、Watanabe N、Matsuda K、Kawata M、Kubo T：“使用转基因大鼠进行骨软骨缺损再生过程中移植的骨髓来源间充质细胞团的行为”骨科研究会汇刊。

大島康史, 高井信朗, 渡邉信佳, 松田賢一, 河田光博, 久保俊一: "骨髄間葉系細胞塊移植による骨軟骨欠損の修復過程-Hanging Drop Culture法を用いて"日本整形外科学会雑誌. 76(8). S1093 (2002)

Yasushi Oshima、Nobuaki Takai、Nobuyoshi Watanabe、Kenichi Matsuda、Mitsuhiro Kawata、Shunichi Kubo：“骨髓间充质细胞团移植修复骨软骨缺损的过程-使用悬滴培养法”日本骨科学会杂志76（8）。 .S1093（2002）

Taniotsu Hashimoto-Gotho, et al.: "Bone mass increase specific to the female in a line of transgenic mice over expressing humnan osteroblast stimulating factor-1"J.Bone Miner.Metab.. (In press). (2004)

Taniotsu Hashimoto-Gotho 等人：“在过度表达人成骨细胞刺激因子-1 的转基因小鼠系中，雌性的骨量增加”J.Bone Miner.Metab..（正在出版）。

Atsushi Tsujimura, Tamotsu Hashimoto-Gotoh: "Future perspective on the osteoporosis gene therapy."Nippon Rinsho. 62. 794-802 (2004)

Atsushi Tsujimura、Tamotsu Hashimoto-Gotoh：“骨质疏松症基因治疗的未来展望。”Nippon Rinsho。

Oshima Y, Watanabe N, Matsuda K, Takenaka N, Kawata M, Takai S: "Behavior of Graft and Host Cells in Underlying Subchondral Bone after Transplantation of Osteochondral Autograft"Transaction of International Cartilage Repair Society. 4. 3605 (2002)

Oshima Y、Watanabe N、Matsuda K、Takenaka N、Kawata M、Takai S：“自体骨软骨移植后软骨下骨中移植物和宿主细胞的行为”国际软骨修复协会交易。