BINDING OF CALCIUM CRYSTALS WITH RENAL CELLS

钙晶体与肾细胞的结合

• 批准号: 6600908
• 负责人: FREDERICK Gary TOBACK
• 金额: $ 10.78万
• 依托单位: UNIVERSITY OF CHICAGO
• 依托单位国家: 美国
• 财政年份: 2002
• 资助国家: 美国
• 起止时间: 2002-07-01 至 2003-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6600908
• 关键词: calcium; clinical research; cyclic AMP; human subject; hydroxyapatites; laboratory rat; membrane proteins; nephrolithiasis; oxalates; pathologic process; prostaglandin E; receptor

Although nephrolithiasis is a common condition, the sequence of events by which crystals are retained in the kidney and form stones is poorly understood. Therefore, additional knowledge is required to identify susceptible patients and formulate new therapeutic strategies to prevent kidney stones which cause pain, hematuria, urinary tract obstruction and infection, and to eliminate the need for expensive procedures such as extracorporeal shock wave lithotripsy or surgery. Our previous studies demonstrate that calcium oxalate monohydrate (COM) crystals bind within seconds to anionic, sialic acid-containing glycoproteins on the apical surface of kidney epithelial cells in culture (employed to model the tubule), suggesting one mechanism whereby crystals could be retained in the kidney. Identification of those molecules on the renal cell surface that mediate crystal binding, and how their expression is modulated, will greatly increase insight into the pathogenesis of nephrolithiasis. Our Preliminary Studies provide evidence that the capacity of renal cells to bind COM crystals is not static, but is regulated by exogenous prostaglandin E (PGE) and intracellular cyclic AMP. Additionally, our recent work utilizing a novel experimental protocol, crystal-affinity chromatography, provides new information about the nature of receptors on the surface of cultured renal cells that bind calcium oxalate crystals, including one apparently novel Cell Surface Protein (CSP) that has been isolated and partially sequenced. Our Specific Aims are to: 1) Define mechanisms by which PGE modulates renal cell crystal affinity by investigating its effect on adhesion of hydroxyapatite (HA) crystals to cells, and its role during crystal adhesion to wounded monolayers; study PGE release in response to renal cell-crystal interactions; study regulation of intracellular cAMP in response to PGE; define the effect of PGE on renal cell protein and glycoprotein synthesis; and define the expression of specific crystal receptor proteins after exposure to PGE. 2) Identify and characterize renal epithelial cell surface glycoprotein receptor(s) for COM and HA crystals; obtain a full-length cDNA clone encoding an apparently novel CSP COM crystal receptor; study regulation of CSP expression in renal cells; define CSP gene and protein distribution in renal and non-renal tissues; and characterize cell surface receptors for other urinary crystals, and in different types of cells. The results will provide new understanding of the mechanisms that control adhesion of urinary crystals to the surface of kidney epithelial cells.

尽管肾结石病是一种常见的疾病，但对肾脏保留在肾脏和形状结石中的事件序列却鲜为人知。因此，需要额外的知识来识别易感患者并制定新的治疗策略，以防止肾结石引起疼痛，血尿，尿路阻塞和感染，并消除对昂贵手术的需求，例如体外冲击波岩石膜状或手术。我们先前的研究表明，草酸钙一水合物（COM）晶体在几秒钟内与培养基肾上皮细胞顶部的阴离子，唾液酸的糖糖蛋白结合（用于模拟肾小管模型），这表明一种机械性可以保留在肾脏中。鉴定介导晶体结合的肾细胞表面上的那些分子，以及如何调节它们的表达，将大大洞悉肾物石病的发病机理。我们的初步研究提供了证据，表明肾细胞结合COM晶体的能力不是静态的，而是受外源前列腺素E（PGE）和细胞内环状AMP的调节。此外，我们使用新的实验方案晶体亲和力色谱法提供了有关结合草酸钙晶体的培养的肾细胞表面上的受体性质的新信息，其中包括一种已被分离并进行部分测序的明显新颖的细胞表面蛋白（CSP）。我们的具体目的是：1）定义PGE通过研究其对细胞羟基磷灰石（HA）晶体对细胞粘附的影响以及在晶体粘附期间对受伤单层的作用来调节肾细胞晶体亲和力的机制；研究PGE响应肾细胞晶体相互作用；研究对PGE的细胞内cAMP的调节；定义PGE对肾细胞蛋白和糖蛋白合成的影响；并定义暴露于PGE后特定晶体受体蛋白的表达。 2）识别和表征COM和HA晶体的肾上皮细胞表面糖蛋白受体；获得编码明显新颖的CSP COM晶体受体的全长cDNA克隆；研究肾细胞中CSP表达的调节；定义肾脏和非肾脏组织中的CSP基因和蛋白质分布；并在其他类型的细胞中表征细胞表面受体。结果将提供对控制泌尿晶体在肾上皮细胞表面粘附的机制的新理解。