STEM CELL THERAPY FOR DISEASES OF BONE IN A MOUSE MODEL

干细胞治疗小鼠模型中的骨疾病

基本信息

批准号：
6944889
负责人：
CHRISTOPHER NIYIBIZI
金额：
$ 30.69万
依托单位：
PENNSYLVANIA STATE UNIV HERSHEY MED CTR
依托单位国家：
美国
项目类别：
财政年份：
2002
资助国家：
美国
起止时间：
2002-09-27 至 2008-08-31
项目状态：
已结题

项目摘要

The focus of the present proposal is to utilize a mouse model of osteogenesis imperfecta (oim) as a model system to evaluate the potential of the bone marrow derived mesenchymal stem cells (BMSCs) to engraft and participate in repair and regeneration of bone. The mouse has a natural occurring mutation that results in non-expression of proa2(I) chains leading to the accumulation of al(I) homotrimers in tissues. The mouse exhibits osteopenia, cortical thinning and easy fracturing and is an excellent model for evaluating the potential of BMSCs as targets for the treatment of genetic and non-genetic diseases of bone. Recent clinical trial by Horwitz et al. using whole marrow in children with a severe form of OI, demonstrated that BMSCs may offer treatment options for O1. Therefore, the hypotheses to be tested are: BMSCs from normal donor mice administered systemically or locally into syngeneic recipient mice will engraft in the bones of the recipient mice, synthesize authentic bone extracellular matrix and contribute to the structural integrity of the host bone. The following specific aims will be used to test these hypotheses: 1) Demonstrate that the cells infused into oim mice will engraft in bone and in fracture sites created in oim mice 2) Demonstrate that the cells which engraft in bone differentiate into osteoblasts and synthesize the authentic bone extracellular matrix and 3) Demonstrate that the cells that engraft in bone contribute to the structural integrity of bone. To accomplish the above aims, BMSCs will be established from femurs and tibiae of normal donor mice and either marked with retroviruses expressing LacZ or GFP genes to aid in cell tracking or unmarked prior to infusion in oim mice. The fate of the infused cells will be tracked by following expression of the marker genes in tissue and by fluorescent in situ hybridization (fish). Differentiation of the transplanted cells into osteoblasts in vivo will be determined by co-localization of osteocalcin and marker genes and also by in situ hybridization. Synthesis of authentic extraceltular matrix by the infused cells will be analyzed by the determination of the presence of type I collagen comprised of 1 and 2 heterotrimers. Structural integrity of the host bone, will be determined by histophotometry, cross-linking, and collagen content and bone mineral density. The proposed studies may lead to the development of better treatments for genetic and non-genetic diseases of bone based on BMSCs.

本提案的重点是利用成骨不全症（oim）小鼠模型作为模型系统来评估骨髓来源的间充质干细胞（BMSC）移植并参与骨修复和再生的潜力。小鼠具有自然发生的突变，导致 proa2(I) 链不表达，从而导致组织中 al(I) 同源三聚体的积累。该小鼠表现出骨质减少、皮质变薄和容易骨折，是评估BMSC作为治疗遗传性和非遗传性骨疾病的靶点潜力的优秀模型。 Horwitz 等人最近进行的临床试验。在患有严重 OI 的儿童中使用全骨髓，证明 BMSC 可以为 O1 提供治疗选择。因此，要检验的假设是：来自正常供体的 BMSC 全身或局部给予同基因受体小鼠的小鼠将植入受体小鼠的骨骼中，合成真实的骨细胞外基质并有助于宿主骨骼的结构完整性。以下具体目标将用于检验这些假设： 1) 证明注入 oim 小鼠体内的细胞将植入到 oim 小鼠的骨骼和骨折部位 2) 证明植入到骨骼中的细胞分化为成骨细胞并合成真实的骨细胞外基质，3) 证明植入骨中的细胞有助于骨的结构完整性。到为了实现上述目标，将从正常供体小鼠的股骨和胫骨中建立 BMSC，并用表达 LacZ 或 GFP 基因的逆转录病毒进行标记以帮助细胞追踪，或者在输注到 oim 小鼠之前不进行标记。输注细胞的命运将通过跟踪组织中标记基因的表达和荧光原位杂交（鱼）来追踪。移植细胞在体内分化成成骨细胞将通过骨钙素和标记基因的共定位以及原位杂交来确定。将通过确定由1和2个异源三聚体组成的I型胶原的存在来分析由输注细胞合成的真实细胞外基质。结构完整性宿主骨将通过组织光度法、交联、胶原蛋白含量和骨矿物质密度来确定。拟议的研究可能会导致基于骨髓间充质干细胞开发更好的遗传性和非遗传性骨疾病治疗方法。