Regulation of pHi and Fluid Flux in Corneal Endothelium

这个项目有三个主要目标：1)证明角膜内皮 泵是基于乳酸盐-水耦合输送的，2)测定详细的生理 乳酸盐-水耦合体系的作用机理；3)开始应用这一基础科学 内皮性营养不良患者逆转角膜水肿的知识。这台泵可以维持角膜 水合作用和透明度。当泵因创伤、炎症、老化或 角膜营养不良，随后出现角膜水肿，透明度丧失，视力显著下降 降级了。通常的治疗方法是移植，这并不是没有重大的妥协。 和并发症。了解泵的工作原理是发展医疗的一种途径 可以推迟或取代移植需求的治疗方法。离子传输是一个关键特征 泵的一部分。在过去的5-7年里，我们提供了强有力的证据表明，泵 工作原理是将乳酸离子从基质输送到前房。这种乳酸盐通量是 由二氧化碳/HCO3-和碳水化合物的细胞和房水缓冲能力促进 无水合酶系统。我们的数据表明，水通量与乳酸通量直接相关。我们 假设内皮细胞产生一种恒定的[乳酸]梯度来驱动水 从基质渗透到前房。其次，我们证明了乳酸的运输是 在内皮营养不良的模型中受到了损害。因此，我们建议提高乳酸盐 通量可以减轻内皮营养不良患者的角膜水肿。使用多个体外、体内和EX 活体互补的方法这将在三个目标中进行测试。AIM#1将提供 填补证据空白的实验条件，这样我们就可以确认假设 由主动转运机制促进的乳酸流量是角膜的主要贡献者。 内皮泵。目标2将通过以下方式确定液体通量与乳酸通量耦合的机制 测试跨内皮细胞[乳酸]站立梯度存在的假设并进行检验 另一种假设认为水通量与单羧酸共转运体直接相关 (MCT)通量。目标3将检验内皮泵功能的假设，在正常和 受压细胞，可以通过低氧适应来增强。如果这一假设是正确的，它将 有潜力作为富氏营养不良症的治疗方法进行测试。

There are three major goals of this project period: 1) to demonstrate that the corneal endothelial PUMP is based on Lactate-Water coupled transport, 2) to determine the detailed physiological mechanism for the Lactate-Water coupled system, and 3) to begin to use this basic science knowledge to reverse corneal edema in Endothelial Dystrophy. The PUMP maintains corneal hydration and transparency. When the PUMP fails due to trauma, inflammation, ageing, or corneal dystrophy, corneal edema ensues, transparency is lost and vision is significantly degraded. The usual therapy is transplantation, which is not without significant compromises and complications. Knowing how the PUMP works is one approach to developing medical therapies that could delay or supplant the need for transplantation. Ion transport is a key feature of the PUMP. Over the previous 5-7 years we have provided strong evidence that the PUMP works by transporting lactate ions from the stroma to the anterior chamber. This lactate flux is facilitated by cellular and aqueous humor buffering power from CO2/HCO3- and the carbonic anhydrase system. Our data indicate that water flux is directly linked with lactate flux. We hypothesize that the endothelial cells create a standing [lactate] gradient that drives water osmotically from the stroma to the anterior chamber. Secondly, we show that lactate transport is compromised in a model of endothelial dystrophy. Therefore, we propose that enhancing lactate flux can reduce corneal edema in endothelial dystrophies. Using multiple in vitro, in vivo & ex vivo complementary approaches this will be tested in three aims. Aim #1 will provide experimental conditions that fill gaps in the evidence so that we can confirm the hypothesis that lactate flux, facilitated by active transport mechanisms, is the major contributor to the corneal endothelial PUMP. Aim 2 will determine the mechanism for coupling fluid flux to lactate flux by testing the hypothesis that a transendothelial [lactate] standing-gradient is present and examine the alternate hypothesis that water flux is linked directly to monocarboxylate cotransporter (MCT) flux. Aim 3 will test the hypothesis that Endothelial PUMP function, in normal and distressed cells, can be enhanced via Hypoxia Adaptation. If this hypothesis is correct, it will have the potential to be tested as a therapy for Fuchs Dystrophy.