Sphingolipids in Acute Kidney Injury

描述(由申请人提供)：研究的总体目标是了解急性肾损伤(AKI)的机制，以便设计新的治疗剂来预防和治疗这种毁灭性的疾病。在过去几年中发展起来的一种策略涉及鞘氨醇1-磷酸受体(S1PR)激动剂。这些化合物消除了与肾脏和其他器官的缺血再灌注损伤(IRI)相关的病理。细胞鞘糖脂在细胞膜受体刺激或质膜损伤后迅速代谢为一系列生物活性中间体，包括1-磷酸鞘氨醇(S1P)。S1P通过细胞表面受体对免疫和非免疫细胞产生不同的细胞信号反应。本研究的重点是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激动剂的保护作用在一定程度上独立于S1P激动剂的典型作用-抑制淋巴细胞转运导致淋巴细胞减少，因为S1P1R激动剂保护缺乏T、B和NKT细胞的RAG-1缺失小鼠的肾脏免受IRI。因此，近曲小管上皮细胞、内皮细胞或髓系细胞(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，缺乏骨髓来源的S1P3Rs可以提供组织保护。这些数据和S1P1Rs和S1P3Rs是肾脏中表达最丰富的S1PR的观察结果支持了目前的观点，即这两种受体亚型在介导组织保护中的外在细胞功能。我们假设：1)S1P1R激活和S1P3R拮抗介导上皮细胞和内皮细胞的组织保护。2)理想的组织保护作用新化合物具有S1P1R激动剂活性和S1P3R拮抗剂活性。提出了三个目标：AIMS 1验证了S1P1R在上皮和内皮细胞上的激活介导近端小管组织保护的假设。目的2验证S1P1R激活和S1P3R缺乏减弱树突状细胞激活IRI天然免疫应答的假说，目的3验证具有S1P1R激动剂活性和S1P3R拮抗剂活性的新化合物是介导组织保护的理想特征的假说。我们将使用野生型和转基因小鼠、嵌合体小鼠、在敏感的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的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执行每个目标。我们的长期计划是更好地了解一类重要受体的作用，这将有助于开发新的治疗干预措施，使用S1P受体化合物来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.