Keratinocyte Galvanotaxis and Wound Healing

描述（由申请人提供）有越来越多的证据表明， 内源性电场在组织修复和重塑中的重要性。我们 这项工作已经将内源性电场的生理相关性扩展到皮肤伤口愈合，表明产生的大小的DC场 在受伤后立即提供角质形成细胞迁移的指导线索。 角质形成细胞在生理条件下的直流电场中向阴极迁移 的中心位置，而在这个位置上的中心位置。 在伤口处，引导角质形成细胞的定向迁移以促进伤口 再上皮化我们现在已经开发了工具和资源来剖析 在伤口愈合的这一重要方面所涉及的机制， 其应用可能导致新的治疗方法。之一 我们发现的亮点是， 表皮生长因子受体（EGFR）激酶激活及其 重新定位到细胞的阴极面。这促使我们提出 两个具体问题。EGF受体是如何定位于表皮细胞的 面对细胞？我们将确定直流电场的应用是否 EGF受体携带囊泡的局部阴极胞吐作用。局部 胞吐作用会在细胞的阴极表面增加膜， 板状Lipodial伸展和定向运动，我们将检查细胞 在电池的阴极和阳极面对端的膜张力来解决这个问题 问题其次，由于趋电反应依赖于EGF受体激酶活性，我们将剖析其上游途径， 在直流电场中局部激活。由于活性氧（ROS）可以 激活EGFR，我们将确定DC场是否产生极化至 阴极细胞膜ROS的上调或下调对细胞凋亡的影响 将研究定向运动性和EGFR磷酸化。此外，委员会认为， 我们的初步数据表明G蛋白偶联β 2肾上腺素能受体 （B2AR），在角质形成细胞迁移中，因此EGFR通过DC的反式激活 将检查B2 AR的场诱导激活。定向调解 通过B2AR与GTP结合Gi或Gs的选择性结合的迁移 蛋白质将被确定。我们认为，多种途径可能会汇聚在 EGFR响应于DC场而启动定向迁移。 解剖控制趋电性的途径可能会提供一个 的基本细胞功能的定向迁移， 趋化性，并将最终指导我们潜在的治疗干预措施 以促进伤口愈合

DESCRIPTION (provided by applicant) There is increasing evidence of the importance of endogenous electric fields in tissue repair and remodeling. Our work has extended the physiological relevance of endogenous electric fields to skin wound healing, demonstrating that DC fields of the magnitude generated immediately upon wounding provide guidance cues for keratinocyte migration. Keratinocytes migrate toward the cathode in DC electric fields of physiological strength, and since the cathodal pole of the field is at the center of the wound, directional migration of keratinocytes is guided to facilitate wound re-epithelialization. We now have developed the tools and resources to dissect the mechanisms involved in this important aspect of wound healing, the application of which may lead to novel therapeutic approaches. One of the highlights of our fmdings has been the demonstration of a requirement for epidermal growth factor receptor (EGFR) kinase activation and its re-localization to the cathodal face of the cell. This prompts us to propose two specific issues. First, how is the EGF receptor localized to the cathodal face of the cell? We will determine if DC electric field application directs localized cathodal exocytosis of EGF receptor-carrying vesicles. Localized exocytosis would add membrane at the cathodal face of the cell, allowing lamellipodial extension and directional movement, and we will examine cell membrane tension at cathodal and anodal facing ends of the cell to address this question. Second, because the galvanotaxis response is dependent on EGF receptor kinase activity, we will dissect the upstream pathways for its localized activation in DC fields. Since reactive oxygen species (ROS) can activate EGFR, we will determine if DC fields generate ROS polarized to the cathodal cell membrane. The effects of up or down-regulation of ROS on directional motility and EGFR phosphorylation will be investigated. Moreover, our pilot data implicate the G protein-coupled beta2-adrenergic receptor (B2AR), in keratinocyte migration, so transactivation of the EGFR through DC field-induced activation of the B2AR will be examined. Mediation of directional migration by selective association of B2AR with either the GTP-binding Gi or Gs proteins will be determined. We submit that multiple pathways may converge upon the EGFR to initiate directional migration in response to the DC field. Dissection of the pathways governing galvanotaxis will likely provide a paradigm for understanding the basic cell function of directional migration in chemotaxis and will ultimately direct us to potential therapeutic interventions to enhance wound healing.