RENAL BASIS OF HYPOCITRATURIA

说明：（直接取自申请表）少尿是肾结石的重要原因。柠檬酸盐在肾小球中自由过滤，尿中柠檬酸盐的排泄主要受近端肾小管中柠檬酸盐重吸收和代谢的速率调节。重吸收由NaDC-1基因编码的顶膜3Na+/柠檬酸共转运体介导，代谢由两条途径之一介导：在三羧酸循环中介导柠檬酸代谢的线粒体途径；以及ATP柠檬酸裂解酶将柠檬酸代谢为乙酰辅酶a和草酰乙酸的细胞质途径。拟开展的研究将探讨调控柠檬酸盐重吸收和代谢的分子机制，重点关注NaDC-1。Aim 1的研究将探讨慢性代谢性酸中毒、钾离子缺乏、碱喂养和饥饿大鼠中NaDC-1调控的分子机制。Aim 2的研究将进一步研究NaDC-1在暴露于酸性细胞外液的培养近端小管细胞中的调控分子机制。这些研究将利用表达天然NaDC-1的细胞，以及表达稳定转染的标记NaDC-1的细胞，以及瞬时转染的报告基因构建物。目标1和目标2将共同讨论NaDC-1的调控。Aim 3的研究将解决已知由酸中毒激活的信号通路在调节NaDC-1中的作用。这些途径包括：1)酪氨酸激酶和MAP激酶途径；2)糖皮质激素;3)内皮素。Aim 4的研究将使用13C核磁共振光谱定量测定肾近端小管线粒体和细胞质途径中的柠檬酸代谢。最后，Aim 5的研究将探讨高钾血症是否会导致高尿症，从而导致高钾血症肾远端小管酸中毒患者缺乏肾结石易感性。这些研究将使我们能够继续了解近端小管中肾柠檬酸盐处理的调节机制。

Description: (Taken directly from the application) Hypocitraturia is an important cause of kidney stones. Citrate is freely filtered in the glomerulus, and urinary citrate excretion is regulated principally by the rate of citrate reabsorption and metabolism in the proximal tubule. Reabsorption is mediated by an apical membrane 3Na+/citrate cotransporter encoded by the NaDC-1 gene, and metabolism is mediate by one of two pathways: a mitochondrial pathway that mediates citrate metabolism in the tricarboxylic acid cycle; and a cytoplasmic pathway in which ATP citrate lyase metabolizes citrate to acetyl CoA and oxaloacetate. The proposed studies will examine the molecular mechanisms responsible for regulation of citrate reabsorption and metabolism, focusing on NaDC-1. Studies in Aim 1 will address the molecular mechanisms of NaDC-1 regulation in rats with chronic metabolic acidosis, K+ deficiency, alkali feeding, and starvation. Studies in Aim 2 will further address the molecular mechanisms of NaDC-1 regulation in cultured proximal tubule cells exposed to acidic extracellular fluid. These studies will utilize cells expressing native NaDC-1, as well as cells expressing stably transfected tagged NaDC-1, and transiently transfected reporter constructs. Aim 1 and 2 together will address the regulation of NaDC-1. Studies in Aim 3 will address the role of signaling pathways known to be activated by acidosis in the regulation of NaDC-1. These pathways include: 1) tyrosine kinase and MAP kinase pathways; 2) glucocorticoids; and 3) endothelin. Studies in Aim 4 will quantitate citrate metabolism in mitochondrial and cytoplasmic pathways of the renal proximal tubule using 13C NMR spectroscopy. Lastly, studies in Aim 5 will address whether hyperkalemia causes hypercitraturia and thus is responsible for the lack of predisposition to nephrolithiasis in hyperkalemic renal distal tubular acidosis. These studies will allow us to continue to pursue an understanding of the mechanisms of regulation of renal citrate handling in the proximal tubule.