Engineering functional anisotropy in chondrocyte- and MSC-laden hydrogels

描述(由申请人提供)： 关节软骨排列在关节表面和功能上，以传递与关节负荷相关的力。受其较差的修复能力的限制，基于细胞的修复策略的需求日益增长。考虑到天然组织的机械作用，使用工程化结构成功地替换关节软骨将需要生长的组织具有功能机械特性。在这项研究中，我们提出了在水凝胶环境(琼脂和自组装多肽水凝胶)中结合软骨细胞或间充质干细胞(可以进行软骨分化)，并重点研究了这些结构的压缩和拉伸性能的发展。成熟软骨的拉伸性能是各向异性的，特别是在表层，随着关节负荷的成熟，在青春期出现。借鉴这一发展概念，我们提出了一种新的生物反应器系统，旨在概括发生在两个接触的关节软骨层之间的滑动接触。使用该生物反应器，将评估MSCs的软骨细胞生物合成和软骨分化作为预孵期和滑动持续时间和间歇性的函数。假设基因表达和分化的模式将由凝胶中的位置与所应用的接触的关系决定，增强发生在接触线的下方和沿线，特别是在表层。后续的长期研究将利用这些优化的加载方案来指导增加MSC和软骨细胞负载的水凝胶结构的压缩和拉伸(沿滑动方向)的机械性能。此外，假设这些结构将显示增加的胶原含量以及在滑动接触方向上平行的胶原纤维取向。这项应用检测了负载软骨细胞和MSC的水凝胶获得软骨样拉伸性能的能力，并通过在新型滑动接触生物反应器系统中的培养将各向异性注入到这些结构中。这些研究将为软骨分化和成熟的机制提供深入的认识。如果实现，这项提案的具体目标将进一步推动我们生产临床相关的软骨细胞和负载MSC的功能性软骨替代物，这种替代物表现出复杂的材料特性和各向异性，这些特性和各向异性定义了天然组织，是其成熟功能所必需的。

DESCRIPTION (provided by applicant): Articular cartilage lines the surfaces of joints and functions to transmit the forces associated with joint loading. Limited by its poor healing capacity, there exists a growing demand for cell-based strategies for repair. Given the mechanical role of the native tissue, the successful replacement of articular cartilage using engineered constructs will require grown tissue possessing functional mechanical properties. In this study, we propose the combination of chondrocytes or mesenchymal stem cells (which can undergo chondrogenic differentiation) in a hydrogel environment (agarose and self-assembling peptide hydrogels), and focus on the development of compressive and tensile properties of these constructs. Tensile properties in mature cartilage are anisotropic, particularly in the superficial zone, and emerge during adolescence as maturation occurs with joint loading. Borrowing from this developmental concept, we suggest a novel bioreactor system designed to recapitulate the sliding contact that occurs between two contacting articular cartilage layers. Using this bioreactor, chondrocyte biosynthesis and chondrogenic differentiation of MSCs will be valuated as a function of pre-incubation period and duration and intermittency of sliding. It is hypothesized that the patterns of gene expression and differentiation will be dictated by the relation of the position in the gel to the applied contact, with enhancements occurring beneath and along the line of contact, particularly in the superficial layer. Subsequent long-term studies will utilize these optimized loading protocols to direct the increase the compressive and tensile (in the direction of sliding) mechanical properties of MSC- and chondrocyte-laden hydrogel constructs. Furthermore, it is hypothesized that these constructs will exhibit enhanced collagen content as well as a parallel collagen fiber orientation in the direction of sliding contact. This application examines the ability of chondrocyte- and MSC-laden hydrogels to achieve cartilage-like tensile properties, and to instill anisotropy in these constructs via culture in a novel sliding contact bioreactor system. These studies will provide insight into the mechanisms of cartilage differentiation and maturation. If realized, the specific aims of this proposal will further our efforts to produce clinically relevant chondrocyteand MSC-laden functional cartilage replacements that exhibit the complex material properties and anisotropies that define the native tissue and are necessary for its mature function.