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.