CaP, crystal responsible for idiopathic nephrolithasis

CaP，导致特发性肾结石的晶体

• 批准号: 7049392
• 负责人: SAEED R. KHAN
• 金额: $ 23.08万
• 依托单位: UNIVERSITY OF FLORIDA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-04-01 至 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7049392
• 关键词: MDCK cell; calcium phosphate; cell adhesion; cell line; clinical research; crystallization; epithelium; human subject; inflammation; nephrolithiasis; oxalates; renal tubule; solutions; urinalysis

DESCRIPTION (provided by applicant): Nephrolithksis is a result of crystal formation and retention within the kidneys. Several studies have shown that conditions are favorable for the formation of calcium phosphate (CaP) in early segments of the nephron namely proximal tubules and the loops of Henle. It is proposed that CaP formed in early segments promote nucleation of calcium oxalate (CaOx) in the collecting ducts. Renal epithelial cells exposed to non-physiologically high levels of oxakte and/or CaOx crystals experience both structural and functional changes, which may eventually lead to cell injury and death. These changes also promote crystal attachment to cell surfaces and retention within the renal tubules. As a protective response, CaOx crystals are often endocytosed at the luminal side and exocytosed into die interstitium where they are processed by the inflammatory defense indicated by the presence of monocytes, macrophages and giant cells. Interestingly CaP crystals seen in the renal interstitium do not induce an inflammatory response. Even though CaP is considered the primary crystal and crystal induced changes in epithelial cells are essential for crystal retention, studies have yet to investigate crystal cell interactions between CaP and renal epithelial cells. CaP has also been shown to dissolve in the acidic environment of the collecting ducts. It is our hypothesis that crystals formed in the urine contain a surface coat, which inhibits crystal dissolution. We also hypothesize that similar to CaOx crystals, an exposure to crystals of CaP also challenges the cells and leads to alterations in renal epithelial cells, which promote crystal retention. We propose to test our hypotheses by exposing proximal tubular origin HK-2 and collecting duct origin MDCK cells to CaP crystals in culture. Two forms of CaP, inorganic apatite and brushite as well as apatite and brushite coated with urinary macromolecules will be used. In addition we propose to investigate the interaction between CaP crystals and renal interstitial fibroblasts to find out why renal interstitial CaP deposits do not cause inflammation. We have the following Specific Aims. 1. To confirm that CaP crystals coated with urinary macromolecules, are resistant to dissolution and good nucleators of CaOx in acidic environment of collecting ducts. 2. To probe that exposure to CaP crystals is associated with renal epithelial cellular injury and that exposure of renal epithelial cells to CaP crystals causes free radical production and lipid peroxidation. 3. To show that production by renal epithelial cells of crystallization modulators such as osteopontin (OPN) and bikunin and a-1 microglobulin is upregulated on exposure to CaP crystals. 4. To demonstrate that CaOx crystal adherence to renal epithelial cells is increased by their previous exposure to CaP crystals and nucleation of CaOx crystals is promoted by surface membranes of the cells pre-exposed to CaP crystals. 5. To determine the molecular mechanism for the inability of renal interstitial CaP crystals to provoke an inflammatory reaction.

描述（由申请人提供）：肾结石是晶体形成和滞留在肾脏内的结果。几项研究表明，条件有利于磷酸钙（CaP）在肾单位的早期部分，即近端小管和Henle袢形成。有人提出，在早期段形成的钙磷促进草酸钙（CaOx）在集合管中的成核。暴露于非生理学高水平的oxakte和/或CaOx晶体的肾上皮细胞经历结构和功能变化，这可能最终导致细胞损伤和死亡。这些变化也促进晶体附着于细胞表面和保留在肾小管内。作为一种保护性反应，CaOx晶体通常在管腔侧被内吞，并被外吞到细胞膜中，在那里它们通过单核细胞、巨噬细胞和巨细胞的存在所指示的炎症防御被加工。有趣的是，在肾间质中观察到的CaP晶体不会诱导炎症反应。尽管CaP被认为是主要的晶体，并且晶体诱导的上皮细胞变化对于晶体保留是必不可少的，但研究尚未调查CaP和肾上皮细胞之间的晶体细胞相互作用。CaP也被证明溶解在集合管的酸性环境中。我们的假设是，尿液中形成的晶体含有表面涂层，可抑制晶体溶解。我们还假设，类似于CaOx晶体，暴露于CaP晶体也会挑战细胞，并导致肾上皮细胞的改变，从而促进晶体保留。我们建议通过将近端肾小管起源HK-2和集合管起源MDCK细胞暴露于培养中的CaP晶体来测试我们的假设。将使用两种形式的CaP，无机磷灰石和透钙磷石以及涂覆有尿大分子的磷灰石和透钙磷石。此外，我们建议研究钙磷晶体和肾间质成纤维细胞之间的相互作用，以找出为什么肾间质钙磷沉积不引起炎症。我们有以下具体目标。1.目的：证实尿大分子包被的CaP晶体在集合管酸性环境中具有抗溶解性和良好的CaOx成核剂。2.目的探讨钙磷晶体与肾上皮细胞损伤的关系，以及钙磷晶体对肾上皮细胞自由基产生和脂质过氧化的影响。3.显示肾上皮细胞的结晶调节剂如骨桥蛋白（OPN）和bikunin和α-1微球蛋白的产生在暴露于CaP晶体时上调。4.为了证明CaOx晶体对肾上皮细胞的粘附通过其先前暴露于CaP晶体而增加，并且CaOx晶体的成核通过预先暴露于CaP晶体的细胞的表面膜而促进。5.确定肾间质CaP晶体不能引起炎症反应的分子机制。