Tissue engineering application of endochondral ossification for bone regeneration

描述（由申请人提供）：目前可用于治疗大节段性缺损的骨移植技术通常会产生血管性差和宿主整合不良的替代组织，导致骨坏死或脱位相关的临床失败。因此，在治疗创伤、骨折不愈合、脊柱融合术、骨坏死和节段性骨截骨术（如肿瘤切除后产生的骨截骨术）的临床结果方面，仍存在显著的未满足需求。组织工程是促进骨再生的一种很有前途的策略。然而，骨组织工程中通过成骨直接刺激骨形成的一般方法在很大程度上是不成功的。在这个建议中，我们通过软骨中间体来实现骨再生。这一过程被称为软骨内成骨，是长骨形成的正常发育机制，也是大多数骨折愈合的途径。我推测通过软骨生成中间体的骨再生将产生一种在形式和功能上都与天然骨相似的新组织。为了验证这一假设，我将比较软骨移植和金标准骨移植技术在一个临界大小的小鼠骨缺损中的骨再生。为了支持这种方法，我有初步的数据表明，含有增生性软骨细胞的软骨移植物促进了血管化和完整的骨再生。除了评估软骨移植与骨移植再生骨的质量外，我还将确定修复发生的机制。根据我的假设，骨再生将通过软骨内成骨过程发生，导致肥大软骨细胞凋亡，产生宿主衍生的骨再生。然而，初步数据表明，供体软骨通过一种尚未解决的机制促进骨再生。我将使用遗传和细胞标记技术来追踪整个修复过程中的细胞表型，以评估软骨如何愈合大骨缺陷。本研究计划的第二个目标是将通过软骨内成骨促进骨再生的概念转化为临床可行的技术。为了实现这一目标，我将设计生物修饰的合成支架，促进间充质干细胞（MSCs）增生软骨的形成。支架将根据软骨内成骨过程设计不同的降解率，以优化体内骨再生。在早期的研究中，我描述了组织工程支架中MSCs的增生性成熟，并开发了一种用于软骨形成的MMP-7生物反应系统。总之，这些目标解决了一个重要的临床问题，可翻译的技术能够改善目前的骨再生技术。此外，这个项目是专门为满足我的长期职业目标而设计的，通过利用多能祖细胞和生物反应支架，在临床相关的体内模型中概括正常发育和/或修复机制。

DESCRIPTION (provided by applicant): Bone graft technologies currently available to treat large segmental defects typically generate replacement tissue with poor vascularity and poor host integration that leads to clinical failures associated with osteonecrosis or dislodgement. As a result, there is a significant unmet need to improve the clinical outcome in procedures treating trauma, fracture non-unions, spinal fusion, osteonecrosis, and segmental bone osteotomies such as those created upon tumor removal. Tissue engineering is a promising strategy to promote bone regeneration. However, the general approach in bone tissue engineering to directly stimulate bone formation through osteogenesis has been largely unsuccessful. In this proposal we approach bone regeneration through a cartilage intermediate. This process, called endochondral ossification, is the normal developmental mechanism for formation of long bones, and is the pathway through which the majority of fractures heal. I hypothesize that bone regeneration through a chondrogenic intermediate will produce a neotissue that resembles the native bone in both form and function. To test this hypothesis I will compare bone regeneration from a cartilage graft to the gold-standard bone graft technique in a critically sized murine bone defect. In support of this approach, I have preliminary data demonstrating that a cartilage graft containing hypertrophic chondrocytes promotes a well-vascularized and integrated bone regenerate. In addition to evaluating the quality of bone regenerated by a cartilage graft versus bone graft, I will determine the mechanism through which repair occurs. According to my hypothesis, bone regeneration will occur through the process of endochondral ossification, resulting in apoptosis of hypertrophic chondrocytes and producing a bone regenerate that is host derived. However, preliminary data indicate that donor cartilage is contributing to the bone regenerate through an unresolved mechanism. I will use genetic and cell labeling techniques to trace the cell phenotype throughout this repair process to evaluate how the cartilage heals large bone defects. The second aim of this research proposal is to translate the concept of promoting bone regeneration through endochondral ossification into a clinically viable technology. To accomplish this I will design biologically modified synthetic scaffolds that promote formation of hypertrophic cartilage from mesenchymal stem cells (MSCs). Scaffold will be designed with variable degradation rates tuned to the process of endochondral ossification to optimize bone regeneration in vivo. In earlier studies I have characterized hypertrophic maturation of MSCs in tissue-engineered scaffolds and developed a MMP-7 bioresponsive system tuned to chondrogenesis. Together these aims address an important clinical problem with a translatable technology capable of improving current bone regeneration techniques. Furthermore this project was specifically designed to meet my long-term career objective related to musculoskeletal regeneration by utilizing multipotent progenitor cells and bioresponsive scaffolds to recapitulate normal development and/or repair mechanisms in clinically relevant in vivo models.