Repair of Focal Defects in the Annulus Fibrosus Using Injectable High-Density Col

描述（由申请人提供）：脊柱中的椎间盘（IVD）由内层髓核（NP）和外层纤维环（AF）组成。腰痛是一种常见的问题，通常通过手术切除突出的椎间盘组织来治疗，在椎间盘组织中留下缺陷。 AF.为了恢复椎间盘的一些功能，已经开发了几种缺损闭合方法，例如缝合和屏障，它们为缺损提供机械支撑，但不促进生物愈合。目前正在开发各种组织工程方法，采用生物材料作为模拟AF结构的支架。这些进展是有前途的，但其在AF缺陷中的性能尚未报告。我们的方法使用体外和体内AF缺损模型，以机械和生物学评价AF修复的生物材料。 AF主要由I型胶原组成，胶原水凝胶已用于许多组织工程应用。作为一种凝胶，胶原蛋白是可注射的，可以很容易地输送到不规则的缺陷;然而，它通常表现出低刚度。我们的实验室已经在全椎间盘置换中使用胶原凝胶，其表现出与周围AF组织的良好整合（Bowles等人，2011年）。因此，我们选择I型胶原蛋白水凝胶作为AF修复的平台。我们将通过三个目标评估胶原蛋白水凝胶用于AF修复的性能，包括体外和体内研究。 第一个目的是评价凝胶密度和交联对胶原凝胶在体外输送至AF缺损处的机械贡献的影响。已经表明胶原蛋白水凝胶的刚度随着胶原蛋白密度而增加（Ibusuki等人，2007年）。同样，与无毒试剂如核黄素交联增加凝胶硬度。我们将筛选具有不同胶原蛋白凝胶密度和核黄素浓度的配方，以找到产生硬凝胶的组合，然后在AF缺损中对该凝胶进行机械测试。 目的2和3侧重于长期AF修复和评估体内缺损模型中的胶原凝胶整合。更具体地说，目标2的目标是评价目标1结果中未接种胶原凝胶保持椎间盘高度和NP含量的能力。当接种绵羊AF细胞时，用于全椎间盘置换的胶原凝胶显示出增加的整合（Bowles等人，2011; Bowles等人，2010年）。因此，目标3将侧重于评估不同细胞密度对凝胶整合的影响， 周围的自然AF组织。当这些研究完成后，将建立评估可注射生物材料的新方法，并表征高密度胶原蛋白用于AF修复的性能。

DESCRIPTION (provided by applicant): Intervertebral discs (IVDs) in the spine consist of an inner nucleus pulposus (NP) and an outer annulus fibrosus (AF). Low back pain is a common problem that is often treated by surgically removing herniated disc tissue, leaving a defect in the AF. In order to return some function to the disc, several defect closure methods have been developed, such as sutures and barriers, which provide mechanical support for the defect but do not promote biological healing. Various tissue engineered approaches are currently being developed that employ biomaterials as scaffolds for AF-mimicking constructs. These advances are promising but their performance in an AF defect has not been reported. Our approach uses both in vitro and in vivo AF defect models to mechanically and biologically evaluate a biomaterial for AF repair. The AF is comprised of primarily type I collagen and collagen hydrogels have been used for many tissue engineering applications. As a gel, collagen is injectable and can be delivered easily to irregular defects; however, it typically exhibits low stiffness. Our lab has used collagen gels in a total disc replacement that exhibited good integration with surrounding AF tissue (Bowles et al., 2011). Therefore we chose type I collagen hydrogels as our platform for AF repair. We will assess the performance of collagen hydrogels for AF repair through three aims encompassing both in vitro and in vivo studies. The first aim will be to evaluate the effects of gel density and crosslinking on the mechanical contribution of collagen gels delivered to AF defects in vitro. It has been shown that the stiffness of collagen hydrogels increases with collagen density (Ibusuki et al., 2007). Likewise, crosslinking with a nontoxic agent like riboflavin increases gel stiffness. We will screen formulations with different collagen gel densities and riboflavin concentrations to find the combination that yields the stiffet gel, then mechanically test that gel in an AF defect. Aims 2 and 3 focus on long term AF repair and assessing collagen gel integration in an in vivo defect model. More specifically, the goal of Aim 2 is to evaluate the ability of unseeded collagen gels taken from Aim 1 outcomes to preserve disc height and NP content. Collagen gels for total disc replacement have shown increased integration when seeded with ovine AF cells (Bowles et al, 2011; Bowles et al., 2010). Therefore Aim 3 will focus on assessing the effect different cell densities on gel integration with surrounding native AF tissue. When these studies are complete, new methods for the assessment of injectable biomaterials will be established and the performance of high-density collagen for AF repair will be characterized.