Regulators of Ischemic Fracture Healing

 描述(申请人提供)：许多骨折表现为受损的再生-愈合缓慢(延迟愈合)或未能愈合(骨不连)。这些损伤可能会因疼痛和功能丧失而复杂化，通常需要进行翻修手术，并且是社会的一大代价。正常的骨再生需要强大的血管反应，因此缺乏血管形成是影响骨愈合的主要危险因素。因此，加强血管形成的方法将促进受损骨折的更大愈合。血栓反应蛋白1(TSP1)及其最亲近的家族成员血栓反应蛋白2(TSP2)是细胞外基质蛋白和强大的内源性血管生成抑制因子，在未愈合的创面中过度表达。TSP1和TSP2在受损骨折愈合的纤维血管阶段高表达，而TSP2的缺失导致缺血性骨折愈合过程中血管形成和骨形成的增强。TSP影响组织血管重建的一个主要机制是这些配体与CD47受体的相互作用，CD47受体是一种强烈影响缺血再灌注的多效性受体。然而，TSP-CD47相互作用在促进缺血性骨折愈合中的意义尚未被研究。我们假设TSP通过与CD47结合，是缺血性骨折愈合的内源性抑制因子。在受损的骨折愈合中，阻断TSP-CD47的结合将促进骨折血管的形成，促进骨再生。为了解决这一假设，我们提出了三个具体目标。首先，我们将通过研究TSP1缺失、TSP2缺失和双基因敲除小鼠来研究TSP1和TSP2在缺血性骨折愈合中的作用。此外，我们还将使用腺病毒重新添加TSP2。在目标2中，我们将探讨CD47是否通过与TSP结合来调节骨折愈合不良时的骨折血管生成和骨再生。我们将检查CD47缺失的小鼠的缺血性骨折愈合，然后评估基于TSP1的CD47激活肽是否抑制缺血性骨折的血运重建。最后，我们将询问在骨折愈合后，使用CD47阻滞剂局部或系统地阻断TSP-CD47配体-受体的相互作用，是否可以在受损的愈合中增强骨折血管形成和骨再生。这些研究的成功完成可能会对折衷骨折患者产生重大的近期临床影响。

 DESCRIPTION (provided by applicant): Many bone fractures show compromised regeneration-healing slowly (delayed union) or failing to heal (non-union). These injuries can be complicated by pain and loss of function, often requiring revision surgery, and are a significant cost to society. A robust vascular response is required for proper bone regeneration, thus a lack of vascularization is a primary risk factor for compromised bone healing. Therefore, methodologies that enhance vascularization will promote greater healing of compromised fractures. Thrombospondin 1 (TSP1) and its closest family member thrombospondin 2 (TSP2) are extracellular matrix proteins and potent endogenous inhibitors of angiogenesis that are over-expressed in non-healing wounds. TSP1 and TSP2 are highly expressed during the fibrovascular phase of compromised fracture healing and an absence of TSP2 results in enhanced vascularization and bone formation during ischemic fracture healing. A dominant mechanism for the influence of TSPs on revascularization in tissues is the interaction of these ligands with the CD47 receptor, a pleiotropic receptor that strongly influences ischemia-reperfusion. However, the significance of TSP-CD47 interactions in enhancing ischemic fracture healing has not been studied. We hypothesize that TSPs are endogenous inhibitors of ischemic fracture healing by binding to CD47. Blocking TSP-CD47 binding will promote fracture vascularization and enhance bone regeneration in compromised fracture healing. To address this hypothesis we propose three specific aims. First we will examine the contribution of TSP1 and TSP2 to ischemic fracture healing by studying TSP1-null, TSP2-null, and double-knockout mice. As well, we will add back TSP2 using adenovirus. In Aim 2, we will ask whether CD47 regulates fracture vascularization and bone regeneration in compromised fracture healing by binding to TSP. We will examine ischemic fracture healing in CD47-null mice, and then evaluate whether a TSP1-based CD47 activating peptide inhibits ischemic fracture revascularization. Finally, we will ask whether disruption of TSP-CD47 ligand-receptor interaction following fracture healing using CD47 blocking agents, delivered either locally or systemically, can enhances fracture vascularization and bone regeneration in compromised healing. Successful completion of these studies could have significant near-term clinical implications for the patient with a compromised fracture.