Sphingolipids in Acute Kidney Injury

描述（由申请人提供）：提出的研究的总体目标是了解急性肾损伤（AKI）的机制，以便设计新的治疗药物来预防和治疗这种破坏性疾病。在过去的几年里，一种发展起来的策略涉及到鞘氨醇1-磷酸受体（S1PR）激动剂。这些化合物消除了与肾和其他器官缺血再灌注损伤（IRI）相关的病理。细胞鞘脂在细胞膜受体受到刺激或细胞膜受到损伤后，迅速代谢为一系列生物活性中间体，包括鞘磷脂1-磷酸（S1P）。S1P通过细胞表面受体对免疫细胞和非免疫细胞产生多种细胞信号反应。本课题主要研究S1P类似物通过S1P1R和S1P3Rs对免疫细胞的细胞外作用，以及肾脏炎症和损伤的细胞机制。我们的初步数据表明，S1P1R激动剂的保护作用部分独立于S1P激动剂的典型作用——抑制淋巴细胞运输导致淋巴细胞减少，因为S1P1R激动剂可以保护缺乏T， B和NKT细胞的rag1缺失小鼠的肾脏免受IRI。因此，近端小管上皮细胞、内皮细胞或髓细胞（树突状细胞）上的S1P1Rs可能是介导组织保护的重要细胞靶点。我们还发现，骨髓来源的S1P3Rs的缺失赋予组织保护。这些数据以及对S1P1Rs和S1P3Rs是肾脏中表达最丰富的s1pr的观察，支持了目前关注这两种受体亚型在介导组织保护中的外源性细胞功能的建议。我们假设：1)S1P1R激活和S1P3R拮抗介导了上皮细胞和内皮细胞的组织保护。2)理想的介导组织保护的新型化合物具有S1P1R的激动剂活性和S1P3R的拮抗剂活性。目的1验证了上皮细胞和内皮细胞上的S1P1R激活介导近端小管组织保护的假设。Aim 2验证了S1P1R激活和S1P3R缺乏会减弱树突状细胞对IRI先天免疫反应的激活的假设，Aim 3验证了具有S1P1R激动剂活性和S1P3Rs拮抗剂活性的新化合物是介导组织保护的理想特征。我们将使用野生型和转基因小鼠、嵌合小鼠、在易感转基因小鼠品系中使用白喉毒素的免疫细胞消融研究、过继转移研究和培养免疫细胞来解决体内和体外的机制问题。此外，我们的方法将结合免疫学、细胞生物学、分子和生理学工具来实现每个目标。我们的长期计划是更好地了解一类重要受体的作用，这将使S1P受体的化合物能够开发新的治疗干预措施，以阻断急性肾损伤的有害后果。

DESCRIPTION (provided by applicant): The overall goal of the research proposed is to understand the mechanisms of acute kidney injury (AKI) to enable the design of new therapeutic agents to prevent and treat this devastating disorder. One strategy that has evolved over the last few years involves sphingosine 1-phosphate receptor (S1PR) agonists. These compounds abrogate the pathology associated with ischemia-reperfusion injury (IRI) of the kidney and other organs. Cellular sphingolipids are metabolized rapidly to a series of bioactive intermediates including sphingosine 1-phosphate (S1P) following stimulation of cell membrane receptors or injury to plasma membranes. S1P has diverse cellular signaling responses on immune and nonimmune cells through cell surface receptors. This proposal focuses on the extracellular effects of S1P analogs through S1P1R and S1P3Rs on immune cells as well as cellular mechanisms of kidney inflammation and injury. Our preliminary data suggest that the protective effect of S1P1R agonists is, in part, independent of S1P agonist's canonical effect - inhibition of lymphocyte trafficking resulting in lymphopenia, because S1P1R agonists protected kidneys from IRI in Rag-1 null mice that lack T, B and NKT cells. Thus S1P1Rs on proximal tubule epithelial cells, endothelial cells or myeloid cells (dendritic cells) could serve as important cellular targets in mediating tissue protection. We also found that the absence of bone marrow derived S1P3Rs confers tissue protection. These data and the observation that S1P1Rs and S1P3Rs are the most abundantly expressed S1PRs in kidney undergird the current proposal focusing on the extrinsic cell function of these two receptor subtypes in mediating tissue protection. We hypothesize that: 1) S1P1R activation and S1P3R antagonism mediate tissue protection in epithelial and endothelial cells. 2) Ideal novel compounds in mediating tissue protection have agonist activity at S1P1R and antagonist activity of S1P3R. Three aims are proposed: Aims 1 tests the hypothesis that S1P1R activation on epithelial and endothelial cells mediate proximal tubule tissue protection. Aim 2 tests the hypothesis that S1P1R activation and S1P3R deficiency attenuates dendritic cell activation of the innate immune response to IRI and Aim 3 tests the hypothesis that new compounds that possess agonist activity at S1P1R and antagonist activity at S1P3Rs are ideal characteristics in mediating tissue protection. We will use wild type and genetically modified mice, chimeric mice, immune cell ablation studies using diptheria toxin in susceptible transgenic mouse strains, adoptive transfer studies, and cultured immune cells to address mechanistic questions both in vivo and in vitro. Furthermore our methods will incorporate immunological, cell biological, molecular and physiological tools in executing each aim. Our long-term plan is to better understand the role of an important class of receptors that that will enable the development of new therapeutic interventions with compounds for S1P receptors to block the deleterious consequences of acute kidney injury. PUBLIC HEALTH RELEVANCE: The incidence and prevalence acute kidney injury (AKI) and its contribution to end-stage renal disease are increasing; mortality is 50 - 80% in some cases. Current treatments have failed in part due the absence of a complete understanding of the underlying pathophysiology. Our studies will identify new pathways that participate in early injury associated with AKI and identify targets for therapeutic intervention using new S1PR- directed compounds.