A New Target for Kidney Stone Prevention: Calcium-Sensitive Transport of Citrate

描述（由申请人提供）：肾结石是一种常见且严重的疾病。尿中柠檬酸盐是钙结石最重要的抑制剂，它能保持钙的可溶性。枸橼酸盐在肾脏中的运输调节已经得到了不充分的调查和仍然知之甚少。一个单一的转运体NaDC1被认为是负责肠和肾柠檬酸重吸收。然而，我们最近发现了一种新的钙敏感的柠檬酸盐运输过程。本应用程序的总体目标是确定肾近端小管通过一种新的根尖钙敏感转运体的根尖柠檬酸盐运输的机制和调节，以及NaDC1和NaDC3的相对作用。我们的中心假设是，当钙增加时，新的柠檬酸盐重吸收过程的作用是限制近端小管中柠檬酸盐的运输。阐明以前未确定的柠檬酸转运蛋白可能会导致新的治疗方法来增加尿中的柠檬酸。目的是：1)确定新的钙敏感柠檬酸转运蛋白及其在近端小管中的作用。基于三个近端小管细胞系的数据，我们的工作假设是，除了NaDC1外，近端小管的顶端侧还存在一种新的钙敏感二羧酸转运蛋白。我们将在这些细胞系中使用NaDC1的敲除来证明这一点。此外，我们将确定柠檬酸盐运输速率和剩余柠檬酸盐运输过程的细胞外钙敏感性

DESCRIPTION (provided by applicant): Kidney stones are a common & serious illness. Urinary citrate is the most important inhibitor of calcium stones by keeping calcium soluble. The regulation of citrate transport in the kidney has received inadequate investigation & remains poorly understood. A single transporter NaDC1 has been assumed to be responsible for intestinal & renal citrate reabsorption. However we have recently identified a novel calcium-sensitive citrate transport process. The overall objective of this application is to determine the mechanisms & regulation of kidney proximal tubule apical citrate transport via a novel apical calcium-sensitive transporter, & the relative roles of NaDC1 & NaDC3. Our central hypothesis is that the novel citrate reabsorptive process functions to limit citrate transport in the proximal tubule when calcium is increased. Elucidation of previously unidentified citrate transporters can potentially lead to new treatments to increase urinary citrate. Aims are: 1) Define the novel calcium-sensitive citrate transporter & its role in the proximal tubule. Our working hypothesis, based on data in three proximal tubule cell lines, is that in addition to NaDC1, a novel calcium- sensitive dicarboxylate transporter is present on the apical side of the proximal tubule. We will use knockdown of NaDC1 in these cell lines to demonstrate this. Also we will determine citrate transport rates & the extracellular calcium-sensitivity for the remaining citrate transport process found in proximal tubule cells grown from dissected proximal tubule segments from knockout mice. 2) Determine the role of NaDC3 in proximal tubule reabsorption of citrate. Based on our surprising preliminary data, we will determine if NaDC3 is expressed on the apical membrane & accounts for calcium- sensitive citrate reabsorption. We will determine the localization of NaDC3 in the proximal tubule & determine the calcium sensitivity on the apical surface. We will also determine the role of basolateral NaDC3. 3) Determine the role of NaDC1 in renal reabsorption & intestinal absorption of citrate. Our working hypothesis is that NaDC1 is not the exclusive mechanism of citrate transport in the kidney & intestine. Other apical mechanisms may both be important in conditions such as acidosis & hypercalciuria. We will use NaDC1 knockout mice & compare renal citrate reabsorption in the -/- to +/+ mice both under baseline & conditions of acidosis & hypercalciuria. We will also determine intestinal citrate absorption in NaDC1 knockout mice.. These studies will establish new paradigms in understanding citrate reabsorption in both the kidney proximal tubule & the intestine, & how these paradigms are impacted in acidosis & hypercalciuria. These outcomes are expected to have an important impact since understanding the regulation of citrate transport in the proximal tubule will dramatically improve strategies in the prevention of calcium stones.