Angiogenesis & Osteogenesis in Structural Graft Healing

描述（由申请人提供）：由于自体皮质骨移植物的供应有限和同种异体结构移植物获得的长期效果较差，修复大型结构性骨缺损仍然是骨科重建手术的主要问题。基于同种异体骨的生物力学和生物相容性，目前还没有结构异体骨移植的替代品，但它们缺乏成骨和骨诱导潜能，再加上宿主无法重塑加工过的皮质骨，导致骨折的比例很高。为了阐明调节结构骨移植愈合的细胞和分子机制，我们开发了小鼠股节段异体移植模型。利用这个模型，我们能够证明自体移植物愈合优于同种异体移植物，因为活体骨膜细胞具有血管生成和成骨活性。我们还开发了在同种异体股骨皮质表面移植细胞和/或重组腺相关病毒（rAAV）的新方法。基于此，我们假设：1)同种异体移植物愈合和后期修复受到血管生成细胞和成骨细胞缺乏的限制。2)利用基于细胞的组织工程方法和/或基于AAV的基因治疗使无生命的同种异体移植物恢复活力，将赋予加工的皮质骨血管生成、成骨和重塑特性，从而增强同种异体骨移植物的愈合。在我们的第一个特异性目标中，我们将利用我们的小鼠股骨骨移植模型进行C57B/6 Rosa 26A雄性小鼠的活体等移植物移植，以正式证明骨膜供体细胞在结构骨移植物血运重建，成骨和重塑中的贡献，在我们的第二个特异性目标中，我们将将骨髓基质细胞植入处理过的同种异体移植物中，以研究其对冷冻同种异体移植物骨愈合和重塑的功效。在我们的第三个目标中，我们将结合干细胞移植和rAAV基因治疗，用活细胞和血管生成-成骨信号补充加工过的结构异体移植物，并比较其与自体移植物和异体移植物的愈合情况。

DESCRIPTION (provided by applicant): Repair of large structural bone defects remains a major problem in orthopaedic reconstruction surgery due to the limited supply of cortical bone autograft and the poor long-term outcomes obtained with structural allografts. While there is no substitute for structural allografts, based on their biomechanical and biocompatible properties, their lack of osteogenic and osteoinductive potential, combined with the host's inability to remodel processed cortical bone, results in a high percentage of fractures. Towards elucidating the cellular and molecular mechanisms that regulate structural bone graft healing, we have developed a murine femoral segmental allograft model. Using this model we were able to demonstrate that autograft healing is superior to that observed with allografts due to the angiogenic and osteogenic activity from live periosteal cells. We have also developed novel approaches to engraft cells and/or recombinant adeno-associated viruses (rAAV) on the cortical surface of the femoral allografts. Based on this, we hypothesize that: 1) allograft healing and later repair is limited by the absence of angiogenic and osteogenic cells. 2) Revitalization of lifeless allograft using a cell based tissue engineering approach and/or AAV based gene therapy will confer angiogenic, osteogenic and remodeling properties on processed cortical bone leading to enhanced bone allograft healing. In our first Specific Aim, we will utilize our murine femoral bone graft model to perform transplantation of live isografts from C57B/6 Rosa 26A male mice to formally demonstrate the contributions of periosteal donor cells in structural bone graft revascularization, osteogenesis and remodeling, in our second Specific Aim, we will seed bone marrow stromal cells onto processed allograft to examine their efficacy on bone healing and remodeling of frozen allograft. In our third aim, we will combine stem cell engraftment and rAAV gene therapy to complement processed structural allograft with live cells and angiogenic-osteogenic signals, and compare its healing with autograft and allograft.