Development of a Multi-phased Scaffold for Soft Tissue to Bone Integration

描述(申请人提供)：前十字韧带(ACL)是最常见的膝关节韧带损伤，在美国每年大约有75,000例韧带手术。自体骨-髌腱-骨移植和肌腱移植是前交叉韧带重建手术中最常用的移植物。在术后早期，骨隧道内的移植物接触部位是力学上最薄弱的部位。因此，前交叉韧带重建手术的临床成功取决于移植物的固定和骨与软组织之间功能性界面的形成。 天然的前交叉韧带-骨附着区由四个不同的区域组成：韧带、非矿化纤维软骨、矿化纤维软骨和骨。我们设计功能界面的方法是通过成骨细胞和韧带成纤维细胞在多相支架系统上进行新型共培养，其结构和功能特性的梯度模仿天然插入区的特性。之所以选择这些设计参数，是因为在整个插入区都观察到了多种类型的单元和矿物含量的固有增加。我们的假设是，成骨细胞和韧带成纤维细胞在具有仿生钙磷(Ca-P)成分的多相支架上共培养将导致支架内形成纤维软骨样界面区。为了检验这一假设，这项提议的具体目标是： 目的1.研究人工前交叉韧带-骨界面的矿物质含量(钙-磷分布、钙/磷比值)。 目的2.建立一种具有仿生钙磷组成变化的多相支架，并检测成骨细胞-韧带成纤维细胞共培养对该支架上界面区特异性标志物(蛋白多糖、II型和X型胶原)的影响。 我们方法的总体假设是，天然组织中的成分和结构分布与功能相关。拟议的对前交叉韧带-骨界面的评估将提供以前无法获得的对插入区组织特征的洞察，并确定具有受控不均质性的支架的设计目标。通过将成骨细胞-成纤维细胞共培养和适当的矿物质含量的带状变化应用到多相支架的设计中，我们试图为开发能够促进愈合并促进体内软组织移植和骨整合的功能性仿生支架奠定基础。

DESCRIPTION (provided by applicant): The anterior cruciate ligament (ACL) is the most frequently injured knee ligament, with approximately 75,000 ligament surgeries performed annually in the United States. The autologous bone-patellar tendon- bone graft and the hamstring tendon graft are the most commonly utilized grafts for ACL reconstruction surgeries. In the early post-operative healing period, the site of graft contact in the bone tunnels represents the weakest point mechanically. Therefore, the clinical success of ACL reconstructive surgeries depends on graft fixation and the formation of a functional interface between bone and soft tissue. The native ACL-bone insertion zone is comprised of four different regions: ligament, non-mineralized fibrocartilage, mineralized fibrocartilage, and bone. Our approach to engineering a functional interface is through the novel co-culture of osteoblasts and ligament fibroblasts on a multi-phased scaffold system with a gradient of structural and functional properties that mimic those of the native insertion zones. These design parameters are chosen because multiple types of cells and an inherent increase in mineral content are both observed across the insertion zone. Our hypothesis is that the co-culture of osteoblasts and ligament fibroblasts on a multi-phased scaffold with a biomimetic calcium phosphate (Ca-P) composition will result in the formation of a fibrocartilage-like interracial zone in the scaffold. To test this hypothesis, the specific aims of this proposal are: Aim 1. To characterize the mineral content (Ca-P distribution, Ca/P ratio) across the native ACL-bone interface. Aim 2. To develop a multi-phased scaffold with a biomimetic compositional variation of Ca-P and to examine the effects of osteoblast-ligament fibroblast co-culture on the development of interfacial zone specific markers (proteoglycan, types II & X collagen) on this scaffold. The global hypothesis of our approach is that the compositional and structural distributions in the native tissue are correlated with functionality. The proposed evaluation of the ACL-bone interface will provide previously unavailable insight into the organizational characteristics at the insertion zone, and identify design objectives for scaffolds with controlled inhomogeneity. By implementing both osteoblast- fibroblast co-culture and the appropriate zonal dependent variations in mineral content into the design of multi-phased scaffolds, we seek to lay the foundation for the development of functional biomimetic scaffolds which can promote healing and facilitate soft tissue graft and bone integration in vivo.