High-Throughput Microenvironment Regulation for Chondrogenesis

 描述(由申请人提供)：肌肉骨骼组织损伤仍然是骨科研究中的一个重大挑战。例如，目前有数百万患者遭受软骨损伤，每年相关的经济成本超过1000亿美元。有几种可行的临床选择来解决这些损伤。在这种背景下，人骨髓间充质干细胞(HMSCs)能够向下分化为软骨形成途径，可以微创的方式从骨髓中获得，并且易于在培养条件下生长，因此是一种很有前途的软骨组织工程细胞来源。尽管hMSCs的分化受可溶性分子、不可溶生化信号和机械信号的调控，但机械负荷和生物材料信号的组合效应在很大程度上是未知的。我们的中心假设是，利用机械刺激下的高通量系统可以阐明单个和协同的微环境因素，以指导hMSCs的软骨分化，从而在体外和在临床相关的体内模型中形成功能性工程软骨。从HTS获得的信息将有助于开发具有增强软骨生成功能的大型结构，这些结构的性能将通过治疗临界大小的关节软骨缺陷在体内得到验证。这些目标将通过实现以下特定目标来实现：(1)开发由不同细胞外基质分子和生长因子组成的三维(3D)组合HTS水凝胶微阵列，其可以机械变形以模拟hMSCs的软骨微环境，(2)定量评估三维组合HTS微阵列和从这些微阵列中选择的大尺度构建物中hMSCs的软骨分化反应，以及(3)确定具有HTS鉴定的成分和机械刺激机制的hMSC构建物在体内诱导软骨再生的潜力。骨科社区将受益于更好地了解这些软骨诱导微环境，最终将诱导新组织形成，并将代表一种可行的替代目前的临床治疗。虽然这项研究的最终目标是设计出临床相关的关节软骨疗法，但这种高温超导技术也可以适用于测试 其他组织的再生策略。

 DESCRIPTION (provided by applicant): Musculoskeletal tissue injuries remain a significant challenge in orthopaedics research. For example, currently, millions of patients are suffering from cartilage injuries, with associated annual financial costs of more than $100 billion dollars. There are several viable clinical options to address these injuries. In this context, human mesenchymal stem cells (hMSCs) are a promising cell source for cartilage tissue engineering as they are capable of differentiating down the chondrogenic pathway, can be obtained from bone marrow in a minimally invasive manner, and are easily grown in culture. Although differentiation of hMSCs is regulated by soluble molecules, insoluble biochemical signals and mechanical cues, the combinatorial effects of mechanical loading and biomaterial signals are largely unknown. Our central hypothesis is that the use of high-throughput systems (HTSs) under mechanical stimulation can be used to elucidate single and synergistic microenvironmental factors for directing the chondrogenic differentiation of hMSCs, and thereby functional engineered cartilage formation in vitro and in a clinically relevant in vivo model. The information obtained from the HTS will help in the development of macroscale constructs with enhanced chondrogenesis, and the performance of these constructs will be validated in vivo by treating critical-sized articular cartilage defects. These goals will be accomplished by achieving the following specific Aims: (1) to develop three dimensional (3D) combinatorial HTS hydrogel-based microarrays, consisting of different extracellular matrix molecules and growth factors, which can be mechanically deformed to mimic the chondrogenic microenvironment of hMSCs, (2) to evaluate quantitatively the chondrogenic differentiation response of hMSCs in 3D combinatorial HTS microarrays and macroscale constructs selected from these microarrays, and (3) to determine the potential of hMSC-laden constructs with HTS-identified compositions and mechanical stimulation regimes to induce cartilage regeneration in vivo. The orthopedic community would benefit from a better understanding of these chondroinductive microenvironments that will ultimately induce neo-tissue formation and will represent a viable alternative to current clinical therapies. While the ultimate objective of this research is to engineer clinically relevant articular cartilage therapies, this HTS can also be applicable to test regeneration strategies for other tissues.