A Biological Basis for Repair of the ACL

描述(申请人提供)：每年有超过100,000名患者发生前交叉韧带(ACL)断裂。它不能自发愈合。由于手术修复和非手术治疗都不能提供可靠的令人满意的临床结果，前交叉韧带断裂仍然是一个紧迫的骨科问题。本提案中描述的研究旨在通过应用新的生物学方法来改善前交叉韧带的修复。这是基于最近的数据显示，与所接受的观点相反，当在韧带的断端之间插入增强型胶原水凝胶时，断裂的前交叉韧带会产生愈合反应。这种反应的关键是细胞从受损的韧带迁移到水凝胶中。初步数据表明，这些细胞不是ACL细胞的代表，但构成了一个独特的群体，具有间充质干细胞(MSCs)的许多特性。为了达到临床上有用的修复，这些细胞必须分裂，分化成韧带细胞，并形成足够机械强度的基质。本提案中描述的实验使用了体外和体内的猪模型来解决这些基本问题。在具体目标1中，从受损的前交叉韧带中迁移的细胞将从水凝胶中恢复，并通过生物学和免疫学标准进行研究，以确定它们在多大程度上具有MSCs的特性。它们在前交叉韧带中的位置将通过免疫组织化学来确定。在具体目标2中，将确定这些细胞对选定的生长因子子集的反应。这些因子是：成纤维细胞生长因子-2、骨形态发生蛋白-12、骨形态发生蛋白-13和转化生长因子-β1。无论是腺病毒介导的基因转移，还是在可能的情况下，重组蛋白都将在单层培养和体外前交叉韧带损伤与修复的“GAP”模型中进行评估。在具体目标3中，将使用猪的全厚度前交叉韧带断裂模型来确定当使用携带基因的增强水凝胶修复临床相关的缺陷时所能达到的机械强度。MRI、组织学和免疫组织化学也将用于评估愈合的韧带。这些研究将促进我们对损伤的前交叉韧带生物学的了解，并提出新的、基于生物学的愈合方法。

DESCRIPTION (provided by applicant): Rupture of the anterior cruciate ligament (ACL) occurs in over 100,000 patients each year. It cannot heal spontaneously. As neither surgical repair nor non-surgical treatments provide a reliably satisfactory clinical outcome, ACL ruptures remain a pressing orthopaedic problem. The research described in this proposal is directed towards improving the repair of the ACL through the application of novel biological approaches. These are based upon recent data suggesting that, contrary to the received view, the ruptured ACL mounts a healing response when a reinforced collagenous hydrogel is interposed between the severed ends of the ligament. Critical to this response is the migration of cells from the damaged ligament into the hydrogel. Preliminary data suggest that these are not representative of ACL cells as a whole, but constitute a distinct population which shares many of the properties of mesenchymal stem cells (MSCs). To achieve a clinically useful repair, these cells must divide, differentiate into ligament cells and lay down a matrix of sufficient mechanical strength. The experiments described in this proposal address these fundamental issues using in vitro and in vivo porcine models. In Specific Aim 1, the cells that migrate from the damaged ACL will be recovered from the hydrogel and investigated by biological and immunological criteria to determine to what degree they share the properties of MSCs. Their location within the ACL will be determined by immunohistochemistry. In Specific Aim 2, the responses of therse cells to a selected sub-set of growth factors will be determined. These factors are: FGF-2, BMP-12, BMP-13 and TGF-beta1. Both adenovirus- mediated gene transfer and, where possible, recombinant proteins will be evaluated in monolayer culture and in an in vitro "gap" model of ACL injury and repair. In Specific Aim 3, a full thickness, ACL rupture model in the pig will be used to determine the mechanical strength that is achievable when a gene-laden, reinforced hydrogel is used to repair a clinically-relevant defect. MRI, histology and immunohistochemistry will also be used to evaluate the healed ligament. These studies will advance our knowledge of the biology of the injured ACL and suggest novel, biologically-based approaches to healing.