SLC4A11 Mitochondrial Uncoupling and ROS Production in Corneal Endothelium

“SLC4A11线粒体解偶联与角膜内皮ROS的产生” 摘要 SLC4A11基因缺陷导致先天性遗传性内皮营养不良和某些形式的 富克斯营养不良症。这项研究的目的是了解这种膜转运蛋白在正常角膜中的作用。 内皮代谢和SLC4A11缺乏如何导致角膜营养不良。角膜内皮 “Pump”维持角膜的水化和透明度。当“泵”由于创伤，炎症， 老化，或营养不良，随之而来的是角膜水肿，透明度丧失，视力显著下降。和往常一样 治疗方法是移植，这并不是没有重大的妥协和并发症。科内尔的标志 内皮性营养不良是线粒体功能障碍。我们的实验室证明SLC4A11是一种NH3 依赖的生电H+转运体。我们发现谷氨酰胺是由血管内皮细胞主动代谢的。 产生NH3，促进ATP的形成。SLC4a11基因敲除显示明显的角膜水肿，乳酸 积累，改变线粒体生理，和ROS。这些数据导致了压倒一切的假设 角膜内皮细胞主动代谢谷氨酰胺，而SLC4A11的缺失改变了谷氨酰胺 代谢，导致线粒体功能障碍，ROS，并最终导致细胞凋亡。初步数据显示， SLC4A11既是一种质膜蛋白，也是一种线粒体膜蛋白，导致了新的假说 SLC4A11是线粒体解偶联分子。使用多种体外和体内互补方法 假说将在三个目标上进行检验。目标1将确定角膜谷氨酰胺代谢是如何促进的 内皮线粒体。假设Slc4a11是一种对NH3敏感的线粒体解偶联器， 与解偶联蛋白-2和潜在的线粒体缓冲牛磺酸一起促进谷氨酰胺 分解代谢。Aim 2将研究ROS和ROS的来源作为对SLC4a11 KO细胞凋亡的刺激。我们的 假设细胞凋亡是由ROS加速的，ROS是由NH3与AN相互作用而产生的 增强了电子传递链，并通过SLC4A11去偶联而减少。目标3将确定角膜的原因 SLC4a11 KO小鼠的水肿。我们将检验失去slc4a11继而导致下调调控的假设。 促进乳酸运输的关键蛋白质。这项研究的完成将确定SLC4A11和 谷氨酰胺在内皮代谢中的作用；为促进谷氨酰胺代谢的机制提供新的见解 减轻NH3诱导的ROS的产生，这将转移到广泛的谷氨酰胺代谢 并为血管内皮细胞营养不良疗法的发展提供了洞察力。

“SLC4A11 Mitochondrial Uncoupling and ROS Production in Corneal Endothelium” ABSTRACT Defects in the gene SLC4A11 cause Congenital Hereditary Endothelial Dystrophy and some forms of Fuchs Dystrophy. The goal of this study is to understand the role of this membrane transporter in normal corneal endothelial metabolism and how SLC4A11 deficiency leads to Corneal Dystrophy. The corneal endothelial “pump” maintains corneal hydration and transparency. When the “pump” fails due to trauma, inflammation, ageing, or 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. A hallmark of Corneal Endothelial Dystrophies is mitochondrial dysfunction. Our laboratory has shown that SLC4A11 is an NH3 dependent electrogenic H+ transporter. We have found that glutamine is actively metabolized by the endothelium producing NH3 and enhancing ATP formation. Slc4a11 knock out shows significant corneal edema, lactate accumulation, altered mitochondrial physiology, and ROS. These data have led to the overarching hypothesis that corneal endothelium actively metabolize glutamine and that the absence of SLC4A11 alters glutamine metabolism, leading to mitochondrial dysfunction, ROS, and eventual apoptosis. Preliminary data indicate that SLC4A11 is both a plasma membrane and a mitochondrial membrane protein, leading to the novel hypothesis that SLC4A11 is a mitochondrial uncoupler. Using multiple in vitro & in vivo complementary approaches these hypotheses will be tested in three aims. Aim 1 will determine how glutamine metabolism is facilitated in Corneal Endothelial mitochondria. The hypothesis is that Slc4a11 is an NH3 sensitive mitochondrial uncoupler that works in conjunction with Uncoupling Protein-2 and the potential mitochondrial buffer taurine to facilitate Glutamine catabolism. Aim 2 will examine the source of ROS and ROS as a stimulus to Apoptosis in Slc4a11 KO. Our hypothesis is that apoptosis is accelerated by ROS, which is generated by the interaction of NH3 with an energized electron transport chain and reduced by SLC4A11 uncoupling. Aim 3 will identify the cause of corneal edema in Slc4a11 KO Mice. We will test the hypothesis that loss of Slc4a11 secondarily induces downregulation of key proteins that facilitate lactate transport. Completion of this study will establish the role of SLC4A11 and glutamine in endothelial metabolism; provide new insight for mechanisms that facilitate glutamine metabolism yet alleviate NH3 induced ROS production that will be transferable to a wide array of glutamine metabolizing tissues; and provide insight for development of therapies for endothelial dystrophies.