Ion Channels and Pumps: The Machinery of Electric Signaling at Corneal Wounds

描述(申请人提供)：角膜上皮细胞迁移和增殖；重要的是，他们这样做是方向性的，以治愈伤口。生长因子和细胞因子在伤口愈合中起关键作用，可能成为角膜创伤治疗的潜在靶点。我们发现了一个非常不同的因素，即角膜伤口处的自然电场(EFS)，它也激活了细胞内的通路。更重要的是，因为EFS本质上是方向性的，它们定向地激活信号通路，给细胞一个方向性的提示，并引导细胞向伤口方向迁移和分裂，以促进愈合。我们的研究表明，EFS超越了其他被广泛接受的定向信号，如接触抑制释放、伤口空洞、群体压力和趋化作用，以引导细胞在定义的方向上迁移。内源性EFS是如何产生和调控的，目前尚不清楚。在链脲佐菌素(STZ)诱导的1型糖尿病大鼠和角膜伤口愈合不良的Pax6/-突变小鼠中，我们观察到内源性伤口EFS显著减少。我们能否增强内源性创面EFS以促进创面愈合，特别是在难治性和慢性创面？我们的长期目标是阐明电信号可以被用来加速伤口愈合的机制。我们最近观察到，创伤后创面EFS逐渐增加，而浴液中的Cl-或Na离子的替代显著改变了内源性EFS。因此，我们假设，角膜损伤诱导了主动调节的创伤电场，这种电场是由氯离子通道和转运分子控制的特定离子(如氯离子)的通量形成的；操纵氯离子通量可能会增强内源性电场，促进伤口愈合。我们将通过以下具体目标来检验这一假说：目的1.证实创伤电场是对伤害的一种主动反应。目的2.探讨内源性EFS在角膜创伤中的离子机制。目的3.阐明创伤电场的分子机制。这一应用的结果将明确角膜创伤愈合中的活跃电信号，提供创伤愈合中电信号的离子和分子机制，并可能导致利用电信号来促进伤口愈合的新疗法。 公共卫生相关性：持续性角膜上皮缺陷是一个重要的医学问题。我们最近在角膜伤口处发现了一种新的信号机制，即自然产生的电场，它对上皮细胞愈合伤口具有深刻的指导作用。该项目旨在确定控制角膜伤口离子通量的离子和分子机制。通过该项目将获得的科学知识，即细胞如何调节这一基本信号，将开辟一条治疗延迟和不可愈合的角膜伤口以及一般伤口的新途径。

DESCRIPTION (provided by applicant): Corneal epithelial cells migrate and proliferate; importantly they do so directionally, to heal wounds. Growth factors and cytokines play pivotal roles in wound healing and may be potential targets for corneal wound therapies. We have discovered a very different factor, namely naturally-occurring electric fields (EFs) at corneal wounds that also activate intracellular pathways. More significantly, because EFs are intrinsically directional, they activate signaling pathways directionally, giving cells a directional cue and guiding cells to migrate and divide in the direction of the wound to facilitate healing. Our studies have shown that EFs override other well-accepted directional cues such as contact inhibition release, wound void, population pressure and chemotaxis to guide cell migration in a defined direction. How the endogenous EFs are generated and regulated is not known. In streptozotocin (STZ)- induced type 1 diabetes mellitus rats and Pax6+/- mutant mice that have defective corneal wound healing, we observed significantly reduced endogenous wound EFs. Can we enhance the endogenous wound EFs to enhance wound healing, especially in refractory and chronic wounds? Our long-term goal is to elucidate the mechanisms through which electric signals can be exploited to accelerate wound healing. We recently observed that wound EFs increase gradually following injury, and substitution of Cl- or Na+ in the bathing solution significantly alters the endogenous EFs. We thus hypothesize that injury to the cornea induces actively-regulated wound electric fields, which are formed by fluxes of specific ions (e.g. Cl-) that are controlled by Cl- channels and transport molecules; manipulating Cl- flux may enhance endogenous electric fields and wound healing. We will test this hypothesis with the following Specific Aims: Aim 1. To confirm that wound electric fields are an active response to injury. Aim 2. To determine the ionic mechanisms of endogenous EFs at corneal wounds. Aim 3. To elucidate the molecular mechanisms of wound electric fields. The results from this application will define the active electric signaling in corneal wound healing, provide ionic and molecular mechanisms of electric signaling in wound healing, and may lead to novel therapies to improve wound healing exploiting electric signaling. PUBLIC HEALTH RELEVANCE: Persistent corneal epithelial defects pose an important medical problem. We recently discovered a novel signaling mechanism at corneal wounds, namely naturally-occurring electric fields that have profound guidance effects on epithelial cells to heal wounds. This project seeks to determine the ionic and molecular mechanisms controlling ionic fluxes at corneal wounds. The scientific knowledge to be acquired through this project, i.e. how cells regulate this fundamental signal, will open a new avenue to treat delayed and non-healing corneal wounds, and wounds in general.