RENAL CRYSTAL GROWTH INHIBITOR PROTEINS

描述(摘自申请者摘要)：尿结石 据信，在形成之前，肾脏中有晶体成核。 管状流体，这些晶体的生长和聚集，以及随后 肾上皮结晶附着。尿蛋白 可能会影响这些过程，但相互作用的性质 尿蛋白与草酸钙和磷酸盐晶体之间的关系 我只了解了一部分。骨桥蛋白(OPN)是一种糖基化的 在培养中由肾皮质细胞分泌的磷蛋白。它 是一种有效的抑制草酸钙晶体生长的体外药物。操作 已在人类尿液和肾结石基质中发现；其作用 在肾结石形成中的作用尚不清楚。对OPN生产的监管已经 在肾脏中没有被研究；然而，在其他组织中产生 受多种激素和生长因子的调节，包括甲状旁腺素和 1，25(OH)2D3.这些激素对蛋白质的影响是 然而，这很复杂，因为它们可能会改变后翻译的类型 蛋白质以特定于组织的方式进行的修饰。 例如，1，25(OH)2D3刺激非磷酸化产物的产生。 OPN在一个靶组织中的形式，并且磷酸化可能改变OPN的 功能。研究人员已从肾皮质细胞中分离出骨桥蛋白 在原代培养中，并使用了针对蛋白质的抗体来 将骨桥蛋白的产生部位定位于肾脏。这项拨款申请 试图解决几个与OPN在 肾脏。因此：1)使用细胞培养系统，并对晶体进行检测 增长，翻译后修改的性质介绍 将研究肾细胞进入OPN，以及它们对 晶体生长。磷酸化、糖基化和蛋白水解酶的作用 将调查硫酸盐化以及翻译后的 通过裂解C末端的多肽来处理OPN；2) 激素刺激对骨桥蛋白产生的调节将在体外进行研究。 在体内，使用检测荷尔蒙对OPN信息的影响的方法，AS 以及蛋白质的分泌。对翻译后修饰的影响 也将寻求；以及3)将描述一种新的检测方法，即 申请人希望提供一种灵敏的方法来检测尿液的影响 大分子对晶体成核的影响。很可能是异常现象 尿晶体生长抑制物在糖尿病发病机制中的作用 肾结石是肾结石的一种亚型。更好地理解 蛋白质影响液体结晶的方式 相对于钙盐过饱和可能会导致更有效的 对这些患者的治疗，也可能有助于更好地 了解发生在其他部位的病理性钙化。

DESCRIPTION (Adapted from the Applicant's Abstract): Urinary calculus formation is believed to be preceded by crystal nucleation in renal tubular fluid, growth and aggregation of these crystals, and subsequent attachment of the crystals of renal epithelium. Urinary proteins probably influence these processes, but the nature of the interaction between urinary proteins and calcium oxalate and phosphate crystals is only partly understood. Osteopontin (OPN) is a glycosylated phosphoprotein that is secreted by renal cortical cells in culture. It is a potent inhibitor of calcium oxalate crystal growth in vitro. OPN has been found in human urine, as well as in renal stone matrix; its role in renal stone formation is unclear. Regulation of OPN production has not been studied in the kidney; however, in other tissues production is regulated by a variety of hormones and growth factors, including PTH and 1,25(OH)2D3. The effect that these hormones have on the protein are complex, however, as they may change the types of post-translational modification that the protein undergoes, in a tissue-specific manner. For example, 1,25(OH)2D3 stimulates production of a non-phosphorylated form of OPN in one target tissue, and phosphorylation may change OPN's function. The investigator has isolated OPN from renal cortical cells in primary culture, and has used antibodies raised to the protein to localize the sites of OPN production in the kidney. This grant request seeks to address several questions related to the function of OPN in the kidney. Thus: 1) using the cell culture system, and assays for crystal growth, the nature of the post-translational modifications introduced into OPN by renal cells will be studied, as well as their effect on crystal growth. The effect of phosphorylation, glycosylation and sulfation will be investigated, as well as the post-translational processing of OPN by cleavage of a peptide from the C-terminal; 2) regulation of OPN production by hormonal stimuli will be studied in vitro and in vivo, using methods to detect hormonal effects on OPN message, as well as protein secretion. Effects on post-translational modifications will also be sought; and 3) a new assay will be characterized, that the applicant hopes will provide a sensitive way to detect effects of urinary macromolecules on crystal nucleation. It is likely that abnormalities of urinary crystal growth inhibitors contribute to the pathogenesis of renal stones in a subset of stone formers. A better understanding of the ways that proteins influence crystallization in fluids that are supersaturated with respect to calcium salts may lead to more effective therapy for these patients, and may also contribute to a better understanding of pathologic calcification occurring in other sites.