Dental Stem Cells and Scaffold-free Tissue Engineering to Enhance Facial Nerve Regeneration

牙科干细胞和无支架组织工程增强面神经再生

• 批准号: 10453479
• 负责人: Fatima Naz Syed-Picard
• 金额: $ 34.15万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-08-12 至 2022-08-11
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10453479
• 关键词: Action Potentials; Address; Adult; Autologous; Autologous Transplantation; Axon; Cell Differentiation process; Cells; Characteristics; Clinical; Crush Injury; Cues; Data; Defect; Dental; Dental Pulp; Development; Effectiveness; Extracellular Matrix; Facial Nerve Injuries; Facial nerve structure; Facial paralysis; Goals; Histologic; In Vitro; Injection of therapeutic agent; Injury; Measurement; Measures; Methods; Modality; Muscle; Natural regeneration; Nerve; Nerve Regeneration; Neural Crest; Neurites; Neuroglia; Neurons; Outcome; Patients; Phenotype; Population; Quality of life; Rattus; Recovery; Recovery of Function; Schwann Cells; Severities; Site; Solid; Spinal Ganglia; Structure; Supporting Cell; Surgical sutures; System; Testing; Time; Tissue Engineering; Tissues; Tooth structure; axon growth; axon guidance; axon regeneration; functional outcomes; ganglion cell; healing; improved; improved outcome; in vivo; mRNA Expression; motor impairment; myelination; nerve injury; nerve transection; neurotrophic factor; novel; protein expression; regenerative; repaired; scaffold; severe injury; soft tissue; standard of care; stem; stem cells; success; three dimensional structure

The overall goal of this study is to accelerate regeneration and improve functional outcomes in facial nerve injuries using novel scaffold-free dental pulp cell constructs. Facial nerve injuries severely impact patient quality of life by impairing motor function and causing facial paralysis. The severity of facial nerve injury dictates treatment modality where mild injuries are treated with end-to-end suturing, and the treatment of more severe injuries involves replacing damaged tissue with autologous nerve grafts; however, these therapies require a prolonged repair time and full functional recovery is not achieved. This is due in part to diminished Schwann cell (SC) support. SCs, neural crest-derived glial cells, provide neurotrophic factors (NTFs), which are known to promote neuron survival and axon extension. The dental pulp, the soft tissue found at the center of the tooth, contains a population of adult stem/progenitor cells that also endogenously express high levels of NTFs, a characteristic attributed to their neural crest origins. Furthermore, dental pulp cells (DPCs) can be induced to differentiate towards a SC-like phenotype, which further enhances NTF expression. We have developed scaffold-free DPC sheets that can be applied for enhancing nerve regeneration in both mild or severe injuries. For mild injury, DPC sheets can be wrapped around injured nerves treated using standard methods to act as localized NTF delivery systems to enhance outcomes. In more severe injuries, DPC sheets can be coaxed into solid, cylindrical tissues that function as conduits that provide a bioactive alternative to autografts. In the latter, DPCs are induced to generate an aligned ECM; therefore, these conduits provide both trophic cues (NTFs) that promote axon regeneration and guidance cues (aligned ECM) that orient axonal growth. Our preliminary data establishes that DPC sheets express high levels of NTFs sufficient to induce neurite outgrowth in neuronal cells in vitro, and enhance regeneration in rat facial nerve crush injury in vivo. Moreover, when cultured on a micro-grooved substrate, DPC sheets produce an aligned ECM that effectively orients neurite extension in vitro. In Specific Aim 1, we will evaluate if inducing SC differentiation will further enhance the regenerative effects of DPC sheets in vitro. We hypothesize that SC-differentiated DPC sheets will have greater NTF expression and induce greater neurite outgrowth in primary dorsal root ganglion cells in vitro than un-induced DPC sheets. In Specific Aim 2, we will test if wrapping an end-to-end sutured transection nerve injury with a DPC sheet will enhance axon regeneration and functional recovery relative to untreated controls. In Specific Aim 3, DPC cell sheets, comprising an aligned ECM, will be rolled in solid, cylindrical conduits and used to bridge segmental injuries in the rat facial nerve. The ability of these conduits to support axonal regeneration and improve nerve functional recovery will be evaluated. The success of these studies will lead to development of a novel and feasible method to accelerate healing and improve functional recovery of damaged facial nerves addressing the major clinical challenge associated with the current standard of care.

本研究的总体目标是加速面神经再生和改善功能结果 使用新的无支架牙髓细胞构建体的损伤。面神经损伤严重影响患者 通过损害运动功能和引起面瘫来影响生活质量。面神经损伤的严重程度 规定了治疗方式，轻度损伤采用端到端的治疗， 严重的损伤涉及用自体神经移植物替换受损组织;然而，这些疗法 需要延长的修复时间并且不能实现完全的功能恢复。这部分是由于减少了 施万细胞（SC）支持。SC，神经嵴源性胶质细胞，提供神经营养因子（NTFs）， 促进神经元存活和轴突延伸。牙髓，在中心发现的软组织 含有一群成体干/祖细胞，这些细胞也内源性表达高水平的 NTFs，一个特征归因于他们的神经嵴起源。此外，牙髓细胞（DPC）可以是 诱导向SC样表型分化，这进一步增强NTF表达。我们有 开发的无支架DPC片材，可用于增强轻度或中度神经再生， 严重受伤。对于轻度损伤，DPC片材可以包裹在使用标准的 方法作为本地化的NTF输送系统，以提高结果。在更严重的伤害中，DPC片材 可以被诱导到固体，圆柱形组织，作为管道，提供生物活性的替代， 自体移植在后者中，DPC被诱导产生对齐的ECM;因此，这些导管提供了两种功能。 促进轴突再生的营养因子（NTFs）和定向轴突的引导因子（对齐的ECM） 增长我们的初步数据确定DPC片表达高水平的NTFs，足以诱导 在体外神经元细胞中突起生长，并在体内促进大鼠面神经挤压损伤的再生。 此外，当在微槽基底上培养时，DPC片产生对齐的ECM，其有效地 在体外定向神经突起延伸。在具体目标1中，我们将评估诱导SC分化是否会进一步 增强DPC片材的体外再生效果。我们假设SC分化的DPC片层 将有更大的NTF表达，并诱导更多的神经突生长在初级背根神经节细胞中， 比未诱导的DPC片体外。在特定目标2中，我们将测试是否包裹端对端缝合横断 相对于未处理的神经损伤，使用DPC片材的神经损伤将增强轴突再生和功能恢复。 对照在具体目标3中，将包含对齐的ECM的DPC细胞片卷成实心圆柱形， 导管和用于桥接大鼠面神经节段性损伤。这些管道支撑 将评价轴突再生和改善神经功能恢复。这些研究的成功将 导致开发了一种新可行的方法来加速愈合和改善功能恢复， 受损的面部神经，解决与当前护理标准相关的主要临床挑战。