The Role of Sustained Calcium Oscillations in the Wound Healing Response

项目摘要 角膜是眼睛最前面的结构，这使得它容易受到抓伤， 感染和营养不良。事实上，受伤和疾病导致的角膜失明是导致角膜失明的第四大原因。 全球可预防的失明这种情况的唯一治愈性治疗是角膜移植或 移植，但活的眼组织并不容易在世界范围内获得。因此，研究 角膜的正常伤口愈合可以为我们提供开发潜在的新治疗方法的信息， 目标的在损伤时，角膜上皮细胞释放激活嘌呤能受体的核苷酸（例如ATP P2 Y2和P2 X7; P2 Y2是G蛋白偶联受体，而P2 X7是配体门控离子通道。两 这些受体诱导短暂的钙波 愈合过程然而，对钙信号在伤口修复中的长期作用知之甚少。给定 P2 X7和P2 Y2的表达和定位在初始钙波后发生变化，我们推测 这些受体也可能参与伤口愈合过程的后期阶段。我们现在已经 确定了钙传播的长期作用，当被抑制时会减弱愈合过程。这 在初级钙波被诱导后10-30分钟发生的次级反应包括： 一系列的钙振荡，在多个细胞簇内传播，并持续了几个小时。 小时用外核苷酸酶灭活细胞外核苷酸抑制了次级反应， 表明在该过程中需要P2 Y2和P2 X7。与主要反应不同，次要反应 钙反应依赖于细胞-细胞接触，因此被亚融合条件和间隙抑制 连接抑制剂。这些结果表明，创伤后的钙反应比创伤后的钙反应更复杂。 这两种受体都是单独的。本课题的目的是研究嘌呤受体P2 X7 和P2 Y2协调并产生继发性钙反应，以及这种反应如何起作用， 伤口愈合的过程因此，我推测嘌呤能受体P2 X7和P2 Y2 差异介导继发性钙反应，并有助于伤口修复中的多个过程。

PROJECT ABSTRACT The cornea is the most anterior structure of the eye, which makes it susceptible to injury from scratching, infections, and dystrophy. In fact, corneal blindness from injury and disease is the fourth highest cause of preventable blindness globally. The only curative treatment for this condition is corneal transplantation or grafting, but viable eye tissues are not readily available worldwide. Therefore, studying the mechanism of normal wound healing in the cornea could give us information on development of potential novel therapeutic targets. Upon injury, corneal epithelial cells release nucleotides (e.g. ATP) that activate purinergic receptors P2Y2 and P2X7; P2Y2 is a G-protein coupled receptor, while P2X7 is a ligand-gated ion channel. Both of these receptors induce a transient calcium wave that has been established to be a critical step in the wound healing process. However, little is known about the long-term role of calcium signaling in wound repair. Given that the expression and localization of P2X7 and P2Y2 change after the initial calcium wave, we speculated that these receptors may also be involved in the later stages of the wound healing process. We have now identified a long-term role of calcium propagation that when inhibited attenuates the healing process. This secondary response, which occurs 10-30 minutes after the primary calcium wave is induced, consists of a series of calcium oscillations that propagate within multiple clusters of cells, and it was seen to last for several hours. Deactivating extracellular nucleotides with ectonucleotidases inhibited the secondary response, indicating that P2Y2 and P2X7 are required in this process. Unlike the primary response, the secondary calcium response depended on cell-cell contact and thus was inhibited by sub-confluent conditions and gap junction inhibitors. These results demonstrate that the calcium response after wounding is more complex than that of either receptor alone. The goal of my project is to investigate how the purinergic receptors P2X7 and P2Y2 coordinate and produce the secondary calcium response and how that response contributes to the wound healing process. Therefore, I hypothesize that purinergic receptors P2X7 and P2Y2 differentially mediate the secondary calcium response and contribute to multiple processes in wound repair.