Dynamic Fibrous Scaffolds for Repairing Dense Connective Tissues

摘要 肌肉骨骼系统的纤维组织（例如膝关节半月板）受到其内在较差的困扰 治愈能力。在之前的融资周期中，我们开发了包括多光纤在内的支持技术 支架引入各种时间和结构信号来修复半月板组织。我们用过 这些支架用于设计具有与天然组织相似的特性和组织的结构（第一个周期）和 然后开发了支架，通过传递因子来招募局部细胞来增强内源性组织修复 细胞（第二个周期）。此次更新的总体目标是进一步改善内源性半月板修复 通过适当的时间和空间编排因子传递来设计支架，首先（i） 软化损伤部位附近的细胞核（通过暂时减少异染色质含量），然后 (ii) 在修复支架内招募并稳定这些细胞的表型。我们假设交付 这些因素将允许将活的内源细胞从半月板募集到支架上，并且 这些因素的空间控制将改善支架定植，即使支架很厚。我们将聘用 复合支架（在之前的资助周期中开发）提供稳定的纤维部分 （聚己内酯（PCL），提供指导模式和机械稳定性），牺牲纤维部分 （聚环氧乙烷（PEO），定义初始支架孔隙率并提供因子早期释放到 环境），以及工程透明质酸（HA）纤维成分（在数周内降解并释放 因素持续存在）。为了解决我们的假设，第一个目标将利用体外微流体- 研究核软化剂（曲古抑菌素 A）、趋化剂（血小板- 衍生生长因子）和纤维软骨形成因子（转化生长因子-β3）以改变核 力学、细胞募集，并促进细胞迁移至纤维的细胞表型的恢复 半月板组织支架。在第二个目标中，我们将控制整个支架的释放（如 之前）或从内层（新提出）跨越各种支架厚度和释放速率 促进厚脚手架的数量。该目标将使用我们最近开发的皮下注射来实现 半月板组织修复模型。在第三个目标中，支架将被植入尤卡坦半岛的半月板缺损处 小型猪在临床相关缺陷模型中评估其功效。如果成功的话，这些研究和 技术将增进我们对内源性半月板工程支架的使用的理解 修复并为临床转化迈出一步。

Abstract Fibrous tissues of the musculoskeletal system (e.g., the knee meniscus) are plagued by their poor intrinsic healing capacity. In the previous funding cycles, we developed enabling technologies including multi-fiber scaffolds to introduce various temporal and structural signals towards the repair of meniscal tissue. We used these scaffolds to engineer constructs with properties and organization similar to native tissues (1st cycle) and then developed scaffolds to enhance endogenous tissue repair through the delivery of factors to recruit local cells (2nd cycle). The overall objective of this renewal is to further improve endogenous meniscus repair with engineered scaffolds through the appropriate temporal and spatial orchestration of factor delivery, to first (i) soften nuclei in cells (via a temporary reduction in heterochromatin content) near the injury site and then (ii) recruit and stabilize the phenotype of these cells within the repair scaffolds. We hypothesize that the delivery of these factors will permit recruitment of viable endogenous cells from the meniscus to the scaffolds and that the spatial control of these factors will improve scaffold colonization, even with thick scaffolds. We will employ composite scaffolds (developed during the previous funding cycles) that provide a stable fiber fraction (polycaprolactone (PCL), to provide an instructional pattern and mechanical stability), a sacrificial fiber fraction (polyethylene oxide (PEO), to define initial scaffold porosity and provide early release of factors into the environment), and an engineered hyaluronic acid (HA) fiber fraction (that degrades over weeks and releases factors in a sustained fashion). To address our hypotheses, the first Aim will utilize in vitro microfluidic- platforms to investigate the timing and dosing of nuclear-softening (Trichostatin A), chemotactic (platelet- derived growth factor), and fibro-chondrogenic factors (transforming growth factor-β3) to alter nuclear mechanics, cell recruitment, and promote resumption of the cellular phenotype of cells migrating into fibrous scaffolds from meniscal tissue. In the second Aim, we will control release from either the entire scaffold (as before) or from an internal layer (newly proposed) across a variety of scaffold thicknesses and release rates to promote population of thick scaffolds. This Aim will be conducted using our recently developed subcutaneous model of meniscus tissue repair. In the third Aim, scaffolds will be implanted into meniscal defects in Yucatan minipigs to evaluate their efficacy in a clinically relevant defect model. If successful, these studies and technologies will advance our understanding of the use of engineered scaffolds for endogenous meniscus repair and provide a step towards clinical translation.