Defining Corneal Schwann cells in Injury

定义损伤中的角膜雪旺细胞

• 批准号: 10308502
• 负责人: ROYCE MOHAN
• 金额: $ 23.86万
• 依托单位: UNIVERSITY OF CONNECTICUT SCH OF MED/DNT
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-12-01 至 2023-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10308502
• 关键词: Affect; Antibodies; Axon; Blood Vessels; Caliber; Cell Differentiation process; Cell Proliferation; Cells; Confocal Microscopy; Cornea; Corneal Injury; Corneal Stroma; Data; Development; Doxycycline; Drug Targeting; Epithelial; Esthesia; Event; Extracellular Matrix; Extracellular Signal Regulated Kinases; Fibrosis; Foundations; Gene Expression; Genes; Genetic; Grant; Impairment; Injury; Investigation; Keratoplasty; Knowledge; Laboratories; Lead; Learning; Lesion; Mechanics; Mesenchymal; Mitogens; Modeling; Molecular; Mouse Protein; Mus; Myofibroblast; Nature; Nerve; Nerve Endings; Nervous System Trauma; Neuroglia; Operative Surgical Procedures; Oryctolagus cuniculus; Outcome; Pathway interactions; Peripheral; Peripheral Nervous System; Pharmacology; Phenotype; Phosphotransferases; Play; Procedures; Process; Proteins; Proteolipids; RNA Sequence Analysis; Regulation; Reporter; Reporter Genes; Role; Schwann Cells; Sensory; Signal Pathway; Signal Transduction; Site; Smooth Muscle Actin Staining Method; Spinal Cord; Stains; Testing; Therapeutic; Time; Tissue Stains; Tissues; Topical application; Transgenic Mice; Transgenic Organisms; Validation; Vision; axon growth; axon regeneration; axonal degeneration; cell behavior; cell type; drug development; enhanced green fluorescent protein; functional outcomes; genetic approach; improved; in vivo; inhibitor; injury and repair; innovation; neovascularization; nerve damage; nerve injury; neurofibroma; novel; novel strategies; overexpression; prevent; programs; protein biomarkers; reinnervation; remyelination; repaired; restoration; sciatic nerve; small molecule; therapeutic target; transcriptome; vascular injury

The transparent cornea is a highly innervated tissue and sustains significant nerve damage during common procedures such as corneal transplantation and vision correction. Unfortunately, the restoration of corneal sensory function after damage is usually inadequate due to aberrant and poor regeneration of axons. In vascularized tissues, such as the sciatic nerve and spinal cord Schwann cells (SCs) are known to support axonal regeneration after injury. However, little is known how corneal SCs respond to injury or surgical procedures as this cell types has not previously been investigated. Studies performed in sciatic injury models reveal that both the genetic encoded factors of SCs, as well as the extracellular matrix components govern the axonal repair process. The dedifferentiation of SCs into a repair SCs – a transient cell type – that re-differentiates into a terminal SC is a hallmark of SC-driven mechanisms in axonal regeneration. In the cornea, the bulk of sensory axons are unmyelinated axons, except at the limbus where they are myelinated. It is presumed that lesions of nonmyelinating corneal axons also enlist the support of their respective SCs for axonal regeneration, mirroring similar activities of SCs of injured vascularized tissues. This idea has not been formally tested before, as experimental evidence to support or refute this paradigm is lacking. To learn what genes are expressed specifically in corneal SCs, we performed a single cell RNA sequence analysis of the rabbit cornea and identified the corneal SC transcriptome. With cross-species validation of several SC-specific target proteins in mouse corneas and validation of a transgenic mouse line expressing proteolipid protein 1-enhanced green fluorescent protein (Plp1-eGFP), we demonstrated SC-specific reporter gene expression in vivo. In this exploratory R21 grant, we propose two aims. In specific aim 1, we will exploit the Plp1- eGFP reporter transgenic line in a corneal stromal injury model causing nerve severance and investigate corneal SC to myofibroblast differentiation over the course of axonal degeneration and repair. These studies will help define whether corneal SCs differentiate into myofibroblasts and nature of injury that promotes this aberrant phenotype. In specific aim 2, we will investigate the wingless (Wnt) signaling pathway in corneal SCs, as molecular components of this pathway showed differential high expression in SCs compared to other corneal cells. We plan to investigate how modulation of the Wnt inhibitor Dickkopf-1 (Dkk1) governs corneal axonal regeneration after injury and effects on corneal mechanical sensation. Together, these objectives will help to lay down the foundation to identify novel targets for SC-therapeutics towards improvement of corneal axonal growth and sensory impairment.

透明的角膜是高度支配的组织，自杀的神经损伤显着 常见程序，例如角膜移植和视力校正。不幸的是， 损伤后的角膜感觉功能的恢复通常由于异常而造成不足 轴突的再生。在血管化组织中，例如坐骨神经和脊髓施旺 已知细胞（SC）在损伤后支持轴突再生。但是，鲜为人知 角膜SC对损伤或手术程序做出反应，因为这种细胞类型以前尚未 调查。在坐骨神经损伤模型中进行的研究表明，两个遗传编码 SC的因素以及细胞外基质组件控制轴突修复过程。 SCS将SCS的去维修SC（瞬态单元格）重新分化为一个重新分化为 末端SC是轴突再生中SC驱动机制的标志。在角膜中，散装 感觉轴突是不髓鞘的轴突，除了在骨髓的边缘。这是 假定非乳脂角膜轴突的病变也征集了他们各自的支持 轴突再生的SC，反映了受伤的血管组织的SC的相似活性。 这个想法以前尚未进行正式测试，作为支持或反驳的实验证据 缺乏范式。要了解在角膜SC中专门表达的基因，我们进行了 兔角膜的单细胞RNA序列分析并鉴定出角膜SC 转录组。用小鼠中几种SC特异性靶蛋白的跨物种验证 角膜和表达蛋白质蛋白1增强绿色的转基因小鼠系的验证 荧光蛋白（PLP1-EGFP），我们在体内证明了SC特异性报告基因表达。 在这项探索性R21赠款中，我们提出了两个目标。在特定目标1中，我们将利用PLP1- 角膜基质损伤模型中的EGFP报告基因转基因线，导致神经隔离和 在轴突变性过程中研究角膜SC至肌纤维细胞分化 维修。这些研究将有助于定义角膜SC是否分化为肌纤维细胞和 促进这种异常表型的伤害的性质。在特定目标2中，我们将调查 角膜SC中的无翅（Wnt）信号通路，作为该途径的分子成分 与其他角膜细胞相比，SC在SC中的表达差异很高。我们计划调查 Wnt抑制剂Dickkopf-1（DKK1）的调节如何控制角膜轴突再生 损伤和对角膜机械感觉的影响。这些目标在一起将有助于提出 在基础上识别SC-治疗学以改善角膜的新型目标 轴突生长和感觉障碍。