Bioactive Scaffold for TMJ Disc Regeneration by Endogenous Stem/Progenitor Cells

项目摘要 据NIDCR估计，颞下颌关节紊乱病（TMJD）影响超过1000万美国人。 共有80 - 90%的症状性TMJDs患者有内部紊乱（ID），也称为椎间盘 移位，这是高度相关的骨关节炎（OA），可能需要手术治疗。 以前尝试用异体和/或合成移植物置换TMJ盘失败，导致进一步的关节炎。 联合退化因此，再生颞下颌关节盘最近出现作为一种替代方法，以克服 目前TMJ疾病治疗的局限性。在我们之前的研究中，我们开发了解剖学上正确的 3D打印的聚己内酯（PCL）支架具有天然的各向异性微纤维取向。设计 天然样异质纤维软骨，结缔组织生长因子（CTGF;促纤维化因子）和 转化生长因子β 3（TGFβ3;软骨形成因子）在空间上嵌入支架中， 包封在聚乳酸-羟基乙酸共聚物（PLGA）微球中（μS）。CTGF的时空释放 和TGFβ3引导的TMJ syMSCs募集，随后进行空间控制的纤维软骨分化 对兔和小型猪的颞下颌关节再生。尽管在体内的结果是有希望的，我们的CTGF/TGFβ3 μ S包埋的支架在TMJ盘再生中遇到了一些突出的翻译挑战， 包括PLGA降解衍生的酸性环境，这是体内 支架的降解速率，以及过度生理剂量的生长因子的潜在副作用。解决 这些问题，在这里，我们建议开发和验证一种新的药物小分子组合， 以替代掺入生物活性支架中的CTGF和TGFβ3，以改善体内降解速率， 通过我们先进的成像模式与新生组织形成平衡，然后促进 临床前大型动物模型中TMJ盘的再生。我们的初步研究确定了新的小 安全、高效和特异性地促进TMJ纤维软骨分化的分子， 衍生的syMSCs。我们还实现了3D打印TMJ盘中小分子的精确控制递送 支架，采用自组装多结构域肽（MDP）水凝胶作为递送载体。我们也 设计了一种高效可靠的成像模式，能够跟踪体内支架降解， 新组织形成。我们将对小分子和 CTGF/TGFβ3作为对照，在我们的支架中递送，关于局部/组织pH变化、细胞毒性、降解 和组织形成。我们将进行全面的体内研究 以平衡支架降解和组织再生。降解速率将通过应用 表面微孔，支架降解以及纤维软骨再生的体内跟踪将是 通过我们的微创成像模式实现。

Project summary Temporomandibular joint disorders (TMJDs) are estimated to affect over 10 million Americans as per NIDCR. Total 80 - 90% of symptomatic TMJDs patients have internal derangement (ID), also referred to as disc displacement, which is highly associated with osteoarthritis (OA) that may necessitate surgical treatment. Previous attempts to replace the TMJ disc with alloplastic and/or synthetic grafts have failed, resulting in further joint degradation. Thus, regeneration of TMJ disc has recently emerged as an alternative approach to overcome limitations of current treatments for TMJ disorders. In our preceding studies, we developed anatomically correct 3D-printed polycaprolactone (PCL) scaffolds with native-like anisotropic microfiber orientation. To engineer the native-like heterogeneous fibrocartilage, connective tissue growth factor (CTGF; profibrogenic cue) and transforming growth factor beta 3 (TGFβ3; chondrogenic cue) were spatially embedded in the scaffolds as encapsulated in poly(lactic-co-glycolic acids) (PLGA) microspheres (μS). The spatiotemporal release of CTGF and TGFβ3 guided recruitment of TMJ syMSCs, followed by spatially controlled fibrocartilaginous differentiation toward regeneration of TMJ in rabbits and mini-pigs. Despite the promising in vivo outcome, our CTGF/TGFβ3 μS-embedded scaffolds encountered few outstanding translational challenges for TMJ discs regeneration, including PLGA degradation-derived acidic environment, a notable interspecies variance in the in vivo degradation rate of scaffolds, and potential side effect of over-physiological dose of growth factor. To address these issues, here we propose to develop and validate a novel combination of pharmaceutical small molecules to replace CTGF and TGFβ3 as incorporated in bioactive scaffolds, to refine the in vivo degradation rate as balanced with de novo tissue formation through our advanced imaging modality, and then to promote regeneration of TMJ discs in a pre-clinical large animal model. Our preliminary study identified novel small molecules that are safe and highly efficient and specific for promoting fibrocartilaginous differentiation of TMJ- derived syMSCs. We also achieved a precisely controlled delivery of the small molecules in 3D-printed TMJ disc scaffolds by adopting a self-assembling multi-domain peptide (MDP) hydrogel as a delivery vehicle. We also devised a highly efficient and reliable imaging modality that will enable to track in vivo scaffold degradation and new tissue formation. We will perform a comprehensive comparative study between small molecules and CTGF/TGFβ3 as control-delivered in our scaffolds regarding local/tissue pH change, cytotoxicity, degradation and tissue formation in our TMJ disc engineering model in vitro. We will conduct a comprehensive in vivo study to balance scaffold degradation with tissue regeneration. The degradation rate will be controlled by applying surface micro-porosity, and in vivo tracking of scaffold degradation as well as fibrocartilage regeneration will be achieved via our minimally invasive imaging modality.