Regulation of pHi and Fluid Flux in Corneal Endothelium

角膜内皮 pHi 和液体流量的调节

• 批准号: 9306379
• 负责人: Joseph Aurelio Bonanno
• 金额: $ 42.68万
• 依托单位: TRUSTEES OF INDIANA UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1991
• 资助国家: 美国
• 起止时间: 1991-07-01 至 2021-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9306379
• 关键词: Active Biological Transport; Address; Aging; Apical; Aqueous Humor; Basic Science; Bicarbonates; Buffers; Carbon Dioxide; Cell Survival; Cell Volumes; Cells; Chimeric Proteins; Cornea; Corneal Endothelium; Corneal dystrophy; Corneal edema; Coupled; Coupling; Data; Distress; Drops; Endothelial Cells; Endothelium; Engineering; Fluorescence Resonance Energy Transfer; Fuchs' Endothelial Dystrophy; Glucose; Goals; HIF1A gene; Human; Hydration status; Hypoxia; Hypoxia Inducible Factor; In Vitro; Inflammation; Ion Transport; Ions; Keratoplasty; Knockout Mice; Knowledge; Lactic acid; Link; Liquid substance; Measures; Medical; Membrane; Mitochondria; Modeling; Mus; Na(+)-K(+)-Exchanging ATPase; Oryctolagus cuniculus; Pathway interactions; Patients; Perfusion; Permeability; Physiological; Production; Property; Protein Isoforms; Pump; Reactive Oxygen Species; Regulation; Reporter; Running; Site; Subfamily lentivirinae; System; Testing; Transplantation; Trauma; Vision; Water; Work; anterior chamber; base; carbonate dehydratase; corneal epithelium; design; experimental study; in vitro testing; in vivo; knock-down; mitochondrial dysfunction; monolayer; sensor; small hairpin RNA; therapy development

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.

本项目的主要目标有三个：1）证明角膜内皮细胞 以乳酸-水耦合运输为基础，2）确定了详细的生理 乳酸-水耦合系统的机制，以及3）开始使用这一基础科学 逆转内皮营养不良症角膜水肿的知识。角膜基质维持角膜 水化和透明度。当由于创伤、炎症、衰老或其他原因导致心脏衰竭时， 角膜营养不良、角膜水肿、透明性丧失和视力显著降低 退化了通常的治疗方法是移植，这不是没有重大的妥协 和并发症。了解神经网络的工作原理是发展医学的一种方法， 可以延迟或取代移植需求的疗法。离子传输是一个关键特征 的。在过去的5-7年里，我们提供了强有力的证据， 通过将乳酸离子从基质输送到前房来起作用。这种乳酸通量是 通过细胞和水状体对CO2/HCO 3-的缓冲能力和二氧化碳的吸收， 脱水酶系统我们的数据表明，水通量与乳酸通量直接相关。我们 假设内皮细胞产生了一个固定的乳酸梯度， 从间质到前房的血管。其次，我们表明，乳酸转运是 在内皮营养不良模型中受损。因此，我们建议， 通量可以减少内皮营养不良中的角膜水肿。使用多种体外、体内和体外 体内补充方法这将在三个目标中进行测试。目标#1将提供 填补证据空白的实验条件，这样我们就可以证实这一假设， 由主动转运机制促进的乳酸通量是角膜上皮细胞的主要贡献者， 内皮细胞目标2将通过以下方式确定耦合液体通量与乳酸通量的机制： 检验存在跨内皮[乳酸盐]站立梯度的假设，并检查 另一种假设是，水通量直接与单羧酸协同转运蛋白有关 (MCT)通量目标3将测试内皮泵功能正常且 受损的细胞，可以通过低氧适应来增强。如果这个假设是正确的， 有可能被测试作为治疗富克斯营养不良症。