Importance of immune-cell lipid signaling in events leading to type 1 diabetes

1型糖尿病（T1D）是胰岛β细胞自身免疫破坏的结果，这一过程的潜在原因尚不完全清楚。我们的研究表明，巨噬细胞和CD4+ t细胞提供的新型脂质信号会影响T1D的发病率。特别是，相关的脂质似乎是由Ca2+独立磷脂酶A2β (iPLA2β)产生的，它在免疫细胞中表达。iPLA2β在sn-2位置水解膜磷脂，释放溶血磷脂和脂肪酸。当脂肪酸是花生四烯酸时，它可以被代谢生成具有生物活性的氧化脂质或类二十烷酸，其中许多是促炎的。我们发现(a) iPLA2β选择性抑制剂，当给药于自发性糖尿病易发非肥胖糖尿病（NOD）小鼠时，可以保持β细胞质量，减少T1D发病率和胰岛素炎，(b) iPLA2β活性促进M1巨噬细胞促炎表型和CD4+ t细胞的TNFα产生，(c)巨噬细胞产生的选择性促炎类20蛋白（PGE2，白三烯，12-HETE, DHETs）在NOD中增加，在iPLA2β (NOD)减少的巨噬细胞中减少。iPLA2β-/+), (d) NOD患者T1D发病率降低。iPLA2β-/+, (e) NOD的过继性转移。iPLA2β-/-巨噬细胞或脾细胞可降低T1D发病率并改善葡萄糖耐量。我们假设CD4+/CD8+ t细胞和巨噬细胞产生的ipla2 β衍生脂质（idl）在T1D的发展中起关键作用，并将在以下目标下解决这一问题：测定CD4+/CD8+ t细胞idl对T1D的贡献。我们建议利用WT和ipla2 β缺失NOD的CD4+和CD8+ t细胞制剂来(a)确定t细胞idl对糖尿病诱导的需求，(b)定量t细胞脂质生成并鉴定idl， (c)评估这些选择的idl对胰岛功能和生存的影响。2. 确定巨噬细胞idl对T1D的贡献。我们建议(a)利用WT和iPLA2β缺失NOD的巨噬细胞制剂来确定巨噬细胞idl对糖尿病诱导的时间需求，(b)评估巨噬细胞iPLA2β选择性缺失NOD的糖尿病发展，(c)确定选择巨噬细胞idl对胰岛功能和生存的影响。3. 评估CD4+/CD8+ t细胞或巨噬细胞来源的idl对胰岛iPLA2β的影响。初步结果表明，应激β-细胞可通过NFκB诱导iPLA2β， PGE2可诱导NFκB。我们将测试巨噬细胞和/或CD4+/CD8+ t细胞idl诱导β细胞iPLA2β的可能性，这将参与维持和放大免疫反应。我们将利用过继转移、系统水平的脂质组学、条件敲除、免疫印迹、信息、IF和选择抑制脂质生成酶的方案来解决这些目标。一下R21目标。短期的。鉴定和评估CD4+/CD8+ t细胞和巨噬细胞idl对T1D发展至关重要。包罗万象的。在免疫治疗的背景下，这可能导致针对和防止这些脂质的产生或修改免疫细胞中负责脂质生成酶的策略。

Type 1 diabetes (T1D) is a consequence of autoimmune destruction of pancreatic islet β-cells and the underlying causes for this process are incompletely understood. Our work suggests novel lipid signaling provided by macrophages and CD4+ T-cells impacts T1D incidence. In particular, the relevant lipids appear to be generated by the Ca2+-independent phospholipase A2β (iPLA2β), which is expressed in immune cells. iPLA2β hydrolyzes membrane phospholipids at the sn-2 position to release a lysophospholipid and a fatty acid. When the fatty acid is arachidonic acid, it can be metabolized to generate bioactive oxidized lipids, or eicosanoids, many of which are pro-inflammatory. We find that (a) an iPLA2β-selective inhibitor, when administered to spontaneous diabetes-prone non-obese diabetic (NOD) mice preserves β-cell mass, reduces T1D incidence and insulitis, (b) iPLA2β activity promotes M1 macrophage pro-inflammatory phenotype and TNFα production from CD4+ T-cells, (c) macrophage production of select pro-inflammatory eicosanoids (PGE2, leukotrienes, 12-HETE, DHETs) is increased in NOD and reduced in macrophages with reduced iPLA2β (NOD.iPLA2β-/+), (d) T1D incidence is reduced in NOD.iPLA2β-/+, (e) adoptive transfer of NOD.iPLA2β-/- macrophages or splenocytes decreases T1D incidence and improves glucose tolerance. We hypothesize that iPLA2β-derived lipids (iDLs) produced by CD4+/CD8+ T-cells and macrophages play critical roles in T1D development and will address this under the following Aims: 1. Determine the contribution of CD4+/CD8+ T-cell-iDLs to T1D. We propose to utilize CD4+ and CD8+ T-cell preparations from WT and iPLA2β-deficient NOD to (a) determine the requirement of T-cell iDLs for diabetes induction, (b) quantitate T-cell lipid production and identify the iDLs, (c) assess the impact of these select iDLs on islet function and survival. 2. Determine the contribution of macrophage-iDLs to T1D. We propose to (a) utilize macrophage preparations from WT and iPLA2β-deficient NOD to determine the temporal requirement of macrophage iDLs on diabetes induction, (b) assess diabetes development in NOD with selective deficiency in macrophage iPLA2β, (c) determine the impact of select macrophage iDLs on islet function and survival. 3. Assess the impact of CD4+/CD8+ T-cell-or macrophage-derived iDLs on islet iPLA2β. Preliminary results suggest that iPLA2β is induced in stressed β-cells by NFκB, and PGE2 has been reported to induce NFκB. We will test the possibility that macrophage and/or CD4+/CD8+ T-cell iDLs induce β-cell iPLA2β, which would be expected to participate in maintaining and amplifying immune responses. We will utilize adoptive transfer, systems level lipidomics, conditional knockouts, immunoblotting, message, IF, and select inhibition of lipid-generating enzymes protocols to address these Aims. R21 Goals. Short-term. Identify and evaluate CD4+/CD8+ T-cell and macrophage iDLs critical for T1D development. Overarching. This could lead to strategies to target and prevent generation of those lipids or modify the responsible lipid-generating enzyme in immune cells, in the context of immunotherapy.