Arginine Uptake Mechanisms Regulate Nitric Oxide in the Renal Vasculature

说明(申请人提供)：一氧化氮(NO)通过影响全身血管阻力和液体和电解质的稳态来调节动脉血压和肾功能。尽管对这一课题进行了大量的研究，但在调节肾脏中NO产生方面的重要因素还没有得到明确的定义。我们实验室以前的工作证明了L-精氨酸(L-精氨酸)摄取机制在肾上皮细胞中的重要性。我们实验室令人振奋的新结果表明，细胞摄取作为一氧化氮合酶底物的L-精氨酸，也是肾血管内皮细胞产生一氧化氮的关键调节因子。因此，L-精氨酸摄取也是肾血管阻力的重要调节因素，从而影响肾脏的排泄功能。新的数据将表明，L-精氨酸转运机制对肾血管中NO的产生和NO依赖功能具有重要影响。利用这些新的数据作为理论基础和独特的综合实验方法，我们将确定L-精氨酸转运体在体外和体内调节肾血管功能中的作用，以提供对这一产生NO的生理调节因子的重要性的理解。实验将验证这一普遍假设，即肾血管内皮细胞Y+和Y+L转运蛋白摄取Y+和Y+L精氨酸通过改变体内一氧化氮合酶底物的可获得性来调节NO的产生，在调节肾功能以及液体和电解质平衡方面发挥关键作用。这一假设将在三个具体目标中得到解决。具体目标1将利用独特的细胞和分子技术来确定肾血管摄取L-精氨酸的机制和转运体。目的2将使用新的方法学来确定这些转运蛋白在正常和疾病动物体内和体外对NO产生和NO依赖功能的功能重要性。然后，目标3中的实验将利用基因操作的小鼠来确定细胞内L-精氨酸摄取机制对哪种一氧化氮合酶亚型的影响。这一新的实验方法组合将被用来阐明L-精氨酸转运在调节肾血管阻力和肾功能中的重要作用。这些研究的结果可能为了解高血压和血管相关并发症的原因提供重要的见解，这些并发症是美国死亡和发病率的主要原因。

DESCRIPTION (provided by applicant): Nitric oxide (NO) regulates arterial blood pressure and kidney function by influencing systemic vascular resistance and fluid and electrolyte homeostasis. Despite the large amount of research on this subject, the factors important in the regulation of NO production in the kidney have not been clearly defined. Previous work from our laboratory demonstrated the importance of L-Arginine (L-Arg) uptake mechanisms in renal epithelial cells. Exciting new results from our laboratory indicate that cellular uptake of L-Arg, the substrate for NO synthase, is also a critical modulator of NO production in renal vascular endothelial cells. L-Arg uptake is therefore also an important regulator of renal vascular resistance with resulting effects on kidney excretory function. New data will be presented indicating that mechanisms of L-Arg transport have important effects on NO production and NO-dependent function in the renal vasculature. Using these novel data as a rationale and a unique integrative experimental approach, we will define the role of the L-Arg transporters in the regulation of renal vascular function in vitro and in vivo to provide an understanding of the importance of this physiological regulator of NO production. Experiments will test the General Hypothesis that cellular L-arginine uptake by y+ and y+L transporters in endothelial cells of the renal vasculature mediates the production of NO by altering the availability of NOS substrate in vivo and plays a critical role in the regulation of renal function as well as fluid and electrolyte homeostasis. This hypothesis will be addressed in three specific aims. Specific Aim 1 will utilize unique cellular and molecular techniques to identify the L-Arg uptake mechanisms and transporters in renal blood vessels. Aim 2 will employ novel methodology to determine the functional importance of these transporters in the renal vasculature on NO production and NO-dependent function both in vitro and in vivo in normal and diseased animals. Experiments in Aim 3 will then make use of genetically manipulated mice to determine which NOS isoform is affected by cellular L-Arg uptake mechanisms. This novel combination of experimental methods will be used to elucidate the important role of L-Arg transport in the regulation of renal vascular resistance and kidney function. The results of these studies may provide important insight into the causes of hypertension and vascular-related complications that are a leading cause of death and morbidity in the US.