Urolithiasis: Oxalate Interactions with the Renal Cells

描述（由申请人提供）：尿石症是一种多因素疾病，不太可能由单一因素导致整个疾病谱系。尽管如此，草酸盐和草酸钙晶体（com -晶体）与肾上皮细胞的相互作用，以及随之而来的细胞反应是重要的，但对这种疾病的研究知之甚少。信号转导机制,调解的影响草酸和COM-crystals在肾结石肾上皮细胞是至关重要的因素,我们的调查的主要目标。五年资助期内,我们表明,草酸和COM-crystals激活p38激酶途径地图和NF-kB转录因子。我们也观察到,人类肾上皮细胞接触草酸以及COM-crystals导致增加生产il - 6,促炎细胞因子,表明代的炎症信号。然而，这些作用的机制尚不清楚。该研究将评估假设p38 MAP激酶介导NF-kB信号在调解中扮演核心角色的影响草酸和COM-crystals,在调节细胞反应。这个假设是由我们的观察,草酸和COM-crystal曝光导致快速和健壮的激活p38 MAP激酶和NF-kB肾上皮细胞。然而，草酸盐和com晶体引起NF-kB活化的确切顺序尚不清楚。目前的研究有四个目标:(1)阐明草酸分子机制参与/ COM-crystal诱导NF-kB通路的激活;(2)研究NF-kB通路在草酸盐/ COM-晶体作用中的介导作用；(3)评价草酸/ COM-crystals调节的分子机制肾上皮细胞的il - 6水平特别强调p38激酶途径地图的作用,NF-kB转录因子激活;和(4)评估p38 MAP激酶信号传导的作用和NF-kB激活体内动物模型的hyperoxaluric肾结石。了解介导草酸盐和COM晶体在肾上皮细胞中的作用的特定信号传导过程将有助于我们制定抑制这些药物病理作用的策略，防止晶体保留和肾结石形成。我们预计，拟议的研究以及我们的初步数据将确定p38 MAP激酶途径和NF-kB作为预防晶体保留的基于机制的靶标。平移的方法,这些研究的长期目标是定义并建立p38 MAP激酶的有用性和NF-kB抑制剂对人类肾结石的预防和治疗的疾病。公共卫生相关性：肾结石疾病是一种多因素疾病，单一因素不太可能导致所有结石患者的亚群。尽管如此,一个重要的因素在《创世纪》的石头疾病是保留晶体材料在肾小管。因此,了解信号机制,调解的影响草酸和COM-crystals后与肾小管细胞是必不可少的在我们的理解与这种疾病相关的分子事件。我们获得的证据表明p38 MAP激酶激活和NF-kB草酸在肾细胞转录因子在体外细胞培养和COM-crystal交互系统。拟建研究的目标是利用人肾细胞原代培养进一步确定这一途径，并验证“p38 MAP激酶途径介导的NF-kB转录因子通过调节基因表达在介导草酸盐和COM-晶体的作用中起核心作用”的假设。我们还寻求在目前的石病实验动物模型中扩展这些观察结果。此外,我们将测试的可行性p38激酶和NF-kB地图的药理抑制剂防止草酸和COM-crystal诱导实验动物病理生理事件。综上所述，我们的研究重点是表征p38 MAP激酶途径和NF-kB作为干预尿石症的分子靶点。因此，本研究方案完全满足项目公告中提出的要求。

DESCRIPTION (provided by applicant): Urolithiasis is a multi-factorial disease and it is unlikely that a single factor will be responsible for the entire spectrum of this disorder. Nonetheless, oxalate and calcium-oxalate crystal (COM-crystal) interactions with renal epithelial cells, and the cellular responses that follow are important, but poorly understood subjects in this disorder. Signal transduction mechanisms that mediate the effects of oxalate and COM-crystals in renal epithelial cells are critical determinants in nephrolithiasis and have been the primary focus of our investigation. During the five year funding period, we demonstrated that oxalate and COM-crystals activate p38 MAP kinase pathway and NF-kB transcription factor. We also observed that exposure of human renal epithelial cells to oxalate as well as COM-crystals results in increased production of IL-6, a pro-inflammatory cytokine, indicating generation of inflammatory signals. However, the mechanism of these actions remains unclear. The proposed studies will evaluate the hypothesis that p38 MAP kinase mediated NF-kB signaling plays central role in mediating the effects of oxalate and COM-crystals, and in modulating the cellular responses. This hypothesis is driven by our observations that oxalate and COM-crystal exposure caused a rapid and robust activation of p38 MAP kinase and NF-kB in renal epithelial cells. However, the precise sequence/s through which oxalate and COM-crystals cause NF-kB activation is not understood. The present studies have four objectives: (1) to elucidate the molecular mechanisms involved in oxalate/COM-crystal induced activation of NF-kB pathway; (2) to investigate the roles played by NF-kB pathway in mediating the effects of oxalate/ COM- crystals; (3) to evaluate the molecular mechanisms by which oxalate/ COM-crystals regulates IL-6 levels in renal epithelial cells with special emphasis on the role of p38 MAP kinase pathway and NF-kB transcription factor activation; and (4) to evaluate the role of p38 MAP kinase signaling and NF-kB activation in vivo animal models of hyperoxaluric nephrolithiasis. Understanding the specific signaling processes that mediate the effects of oxalate and COM crystals in renal epithelial cells will help us develop strategies to inhibit the pathologic effects of these agents and prevent crystal retention and renal stone formation. We anticipate that proposed studies together with our preliminary data will identify p38 MAP kinase pathway and NF-kB as a mechanism-based target/s for the prevention of crystal retention. As a translational approach, the long-range goal of these studies is to define and establish the usefulness of p38 MAP kinase and NF-kB inhibitors for the prevention and therapy of human kidney stone disease.PUBLIC HEALTH RELEVANCE: Kidney stone disease is a multifactorial disorder and it is unlikely that a single factor will be responsible for all the subsets of stone patients. Nonetheless, one important factor in genesis of stone disease is retention of crystalline materials in the renal tubules. Therefore, understanding the signaling mechanisms that mediate the effects of oxalate and COM-crystals following their interaction with renal tubular cells are essential in our understanding the molecular events associated with this disorder. We have obtained evidence indicating activation of p38 MAP kinase and NF-kB transcription factor in renal cells upon oxalate and COM-crystal interaction in vitro cell culture systems. The goal of the proposed studies is to further define this pathway using primary cultures of human kidney cells and to test the hypothesis that, "p38 MAP kinase pathway mediated NF-kB transcription factor plays a central role in mediating the effects of oxalate and COM- crystals by regulating gene expression". We also seek to extend these observations in current experimental animal models of the stone disease. Moreover, we will test feasibility of a pharmacological inhibitors of p38 MAP kinase and NF-kB in preventing oxalate and COM-crystal induced pathophysiological events in the experimental animals. Taken together our studies focus on characterizing p38 MAP kinase pathway and NF-kB as a molecular targets for intervention in urolithiasis. Thus this proposed study completely satisfies the requirements set forth in the Program Announcement.