Multicellular regulation of corneal wound healing

角膜伤口愈合的多细胞调节

基本信息

批准号：
8975202
负责人：
Vickery E Trinkaus-Randall
金额：
$ 20.46万
依托单位：
BOSTON UNIVERSITY MEDICAL CAMPUS
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-12-01 至 2017-11-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8975202
关键词：
Agonist Apyrase Axon Blindness Chemical Injury Coculture Techniques Communication Cornea Corneal Injury Corneal Ulcer Data Disease Ensure Epithelial Epithelial Cells Epithelium Event Exhibits Family Fluorescent Probes Glutamates Goals Health Homeostasis Image In Situ Infection Injury Keratopathy Lead Mediating Membrane Modeling Monitor N-Methyl-D-Aspartate Receptors N-Methylaspartate Natural regeneration Nerve Operative Surgical Procedures Pain Pathology Pattern Phase Photophobia Play Purinoceptor Rattus Recurrence Regulation Role Sensory Signal Pathway Signal Transduction Signaling Molecule Signaling Protein Stress System Therapeutic Time Tissues Trauma Vision Wound Healing Writing afferent nerve cell motility cell type corneal epithelium design eye dryness in vivo in vivo imaging inhibitor/antagonist intercellular communication migration nerve supply prevent receptor repaired research study response response to injury

项目摘要

DESCRIPTION (provided by applicant): The cornea is the most densely innervated tissue in the body. Damage to corneal nerves in the epithelium can originate from trauma including surgery, disease, infection and chemical injury and can lead to dry eye, a keratopathy, that is exhibited by photophobia and pain, corneal ulcers, and, ultimately, blindness. In all of these conditions, signaling molecules are released by both corneal epithelium and nerves, creating potential for auto- and cross-stimulation. However, relatively little is known about this crosstalk during homeostasis or regeneration. To evaluate these potential interactions, we developed a co-culture of nerves and epithelia. Using this model, we demonstrated that Ca2+ mobilization occurs after stimulation of either cell type. Both epithelia and nerves release ATP in response to injury or agonist stimulation and activate a family of purinoreceptors that play a role in Ca2+ mobilization, cell migration, and pain. The mobilization is divided into two phases with an initial rapid increase and decrease followed by a second set of slower oscillations. Pretreatment with apyrase, an ectoenzyme, removed the initial response, whereas pretreatment with N-methyl-D-aspartate (NMDA) inhibitors prevented the second response. Nerves release glutamate, and both epithelia and nerves express NMDA receptors. Preliminary data indicate that environmental stress causes a decrease in the Ca2+ oscillations, a change in localization of the NMDA receptor away from the membrane, and a change in cell-cell signaling. Analyses using MATLAB demonstrate distinct patterns in cell-cell signaling. Since our co-culture system does not capture the complexity of the native connections that exist between sensory axons and corneal epithelial cells, we used the Thy-1-GFP homozygous rat. Preliminary in situ imaging studies performed ex vivo reveal that we can image cytosolic Ca2+ mobilization and fluorescent sensory nerves. Our goal is to determine mechanisms by which sensory nerves and corneal epithelial cells signal to coordinate the response induced by trauma and its impact on migration. We hypothesize that release of ATP by nerves and epithelium and release of trophic factors by nerves regulates the activation of specific purinergic and glutamatergic receptors that mediate wound repair.

描述（由申请人提供）：角膜是体内神经支配最密集的组织。角膜上皮神经的损伤可能源于外伤，包括手术、疾病、感染和化学损伤，并可能导致干眼症、角膜病（表现为畏光和疼痛）、角膜溃疡，最终导致失明。在所有这些情况下，角膜上皮和神经都会释放信号分子，从而产生自动刺激和交叉刺激的潜力。然而，人们对这种串扰知之甚少在稳态或再生期间。为了评估这些潜在的相互作用，我们开发了神经和上皮细胞的共培养物。使用该模型，我们证明了刺激任一细胞类型后都会发生 Ca2+ 动员。上皮和神经都会响应损伤或激动剂刺激而释放 ATP，并激活在 Ca2+ 动员、细胞迁移和疼痛中发挥作用的嘌呤受体家族。动员工作分为两个阶段快速增加和减少，然后是第二组较慢的振荡。用腺苷三磷酸双磷酸酶（一种胞外酶）预处理消除了最初的反应，而用 N-甲基-D-天冬氨酸（NMDA）抑制剂预处理则阻止了第二次反应。神经释放谷氨酸，上皮细胞和神经都表达 NMDA 受体。初步数据表明，环境压力会导致 Ca2+ 振荡减少、NMDA 受体远离膜的定位发生变化，以及细胞间信号传导发生变化。使用 MATLAB 进行的分析展示了细胞间信号传导的不同模式。由于我们的共培养系统无法捕获感觉轴突和角膜上皮细胞之间存在的天然连接的复杂性，因此我们使用了 Thy-1-GFP 纯合大鼠。体外进行的初步原位成像研究表明，我们可以对胞质 Ca2+ 动员和荧光感觉神经进行成像。我们的目标是确定感觉神经和角膜上皮细胞发出信号以协调创伤引起的反应及其对迁移的影响的机制。我们假设神经和上皮释放 ATP 以及神经释放营养因子调节介导伤口修复的特定嘌呤能和谷氨酸能受体的激活。