Dynamic Fibrous Scaffolds for Engineering Dense Connective Tissues

描述(由申请人提供)： 当内源性修复失败时，就像致密的纤维肌肉骨骼组织(如膝关节半月板)经常发生的那样，必须开发新的战略和使能技术来促进组织再生。虽然许多组织工程疗法已经被开发用于纤维增强组织的工程，但一个持久的限制是构建支架的能力，该支架保持结构完整性和指导适当的组织结构，同时促进整个再生期的细胞渗透。在这项提案中，我们通过生产一种新型的多聚合物复合纳米纤维支架来解决这些限制，这种支架为新生组织的形成提供了三维微图案。这些支架的纤维直径约为天然细胞外基质的数量级，可以通过定义的纤维各向异性来模拟纤维增强组织的结构排列。此外，通过光交联型大分子聚合物库引入聚合物性能的灵活性，当聚合时表现出一系列的机械和降解性能，我们建议通过控制单个聚合物组分的降解来定制支架内的临时气孔形成。我们假设这些纳米纤维多聚合物光交联网将具有反映最硬和最慢降解组分的受控机械性能，并在保持其整体力学性能的同时，显示出空隙中随时间的增加。此外，我们假设，通过牺牲元件的侵蚀，这些支架内的空隙被控制地增加，将促进细胞渗透到多聚物网中，并将导致更均匀的功能组织结构。此外，由于半月板的机械环境对其成熟和动态平衡至关重要，我们假设量身定制的机械预适应方案同样会促进浸润性半月板结构的功能成熟。为此，开发了一种新型的动态加载生物反应器，采用压缩和拉伸相结合的方式来促进组织成熟。这项提议的第一个目标是开发电纺多组分纳米纤维支架的技术，并使用预测的纤维增强复合材料模型将其性能与天然组织进行比较。在第二个目标中，将探索单个和三个聚合物细胞负载的纳米纤维支架中细胞的相互作用和渗透以及最终的力学性能。第三个目标涉及开发一种生物反应器，用于对支架进行预调节，并评估机械刺激对组织形成的短期和长期影响。如果成功，这一创新方法将为受损的纤维增强肌肉骨骼组织的功能性再生提供几项新的使能技术。 公共卫生相关声明(由申请人提供)：该项目开发了一种新型的多聚合物纳米纤维制造系统，以控制聚合物化学和整体支架力学以及随时间的降解，以改善纤维组织工程构建物中的细胞渗透和均匀的组织沉积。如果成功，这种方法将克服肌肉骨骼系统致密结构的组织工程中的一个主要障碍，并提供一种机械的 功能、结构各向异性的3D微图案，用于定向新组织的形成，同时促进完全细胞定植，并在完全溶解后最终替换聚合物结构。这种结合支架制作、机械加载和体内评估的创新方法将有助于开发组织工程疗法，用于纤维增强的肌肉骨骼组织，如膝关节半月板，否则这些组织无法愈合，临床上几乎没有可行的修复策略。

DESCRIPTION (provided by applicant): When endogenous repair fails, as is often the case with dense fibrous musculoskeletal tissues (like the knee meniscus), novel strategies and enabling technologies must be developed to enhance tissue regeneration. While numerous tissue engineering therapies have been developed for the engineering of fiber-reinforced tissues, one persistent limitation is the ability to fabricate scaffolds that maintain structural integrity and direct appropriate tissue architecture, while simultaneously promoting cellular infiltration throughout the regeneration period. In this proposal, we address these limitations with the production of a novel multi-polymer composite nanofibrous scaffold that provides a 3-dimensional micropattern for neo-tissue formation. These scaffolds, with fiber diameters on the order of the native extracellular matrix, can be produced with defined fiber anisotropies that mimic the structural arrangement of fiber-reinforced tissues. Further, by introducing flexibility in polymer properties via a library of photocrosslinkable macromers that exhibit a range of mechanical and degradation properties when polymerized, we propose to tailor temporal pore formation within the scaffold through the controlled degradation of individual polymer components. We hypothesize that these nanofibrous multipolymer photocrosslinked meshes will have controlled mechanical properties reflective of the stiffest and slowest degrading component and show a time dependent increase in void space while maintaining their overall mechanical properties. Further, we hypothesize that the controlled increase in void space within these scaffolds, via the erosion of sacrificial elements, will promote cellular infiltration into the multi-polymer mesh and will result in a more uniform functional tissue structure. Additionally, as the mechanical environment of the meniscus is paramount in its maturation and homeostasis, we hypothesize that tailored mechanical preconditioning regimens will likewise promote functional maturation of infiltrated meniscus constructs. To this end, a novel dynamic loading bioreactor that applies compression and tension is developed to promote tissue maturation. The first Aim of this proposal is to develop technology to electrospin multi-component nanofibrous scaffolds and compare properties to the native tissue using a predictive fiber-reinforced composite model. In the second Aim, the interaction and infiltration of cells and ultimate mechanical properties will be explored in single- and tri-polymer cell-laden nanofibrous scaffolds. The third Aim involves the development of a bioreactor for pre-conditioning scaffolds and evaluating the impact of mechanical stimulation on tissue formation in the short and long term. If successful, this innovative approach will provide several new enabling technologies for the functional regeneration of damaged fiber-reinforced musculoskeletal tissues. Public Health Relevance Statement (provided by applicant): This project develops a novel multi-polymer nanofiber fabrication system to exert control over polymer chemistry and overall scaffold mechanics and degradation with time to improve cellular infiltration and uniform tissue deposition in fibrous tissue-engineered constructs. If successful, this approach would surmount a major hurdle in the tissue engineering of dense structures of the musculoskeletal system and provide a mechanically functional, structurally anisotropic 3D micro-pattern for directed neo-tissue formation while promoting full cellular colonization and eventual replacement of the polymer structure after complete dissolution. This innovative approach, coupling scaffold fabrication, mechanical loading, and in vivo assessments, will aid in the development of tissue engineered therapies for fiber-reinforced musculoskeletal tissues such as the knee meniscus that otherwise fail to heal and have few clinically viable repair strategies.