Docosanoids modulate homeostasis and cell survival after ischemic stroke

项目摘要/摘要 尽管神经保护策略显示出了希望，但中风后还没有任何治疗方法证明有效。 本项目重点研究DOC介体的神经保护生物活性：神经保护素D1 (NPD1)、Resolvin D_1(RvD_1)及其组合(NPD1+RvD_1)抗实验性脑缺血和血栓形成 卒中。这些脂质介体是按需生物合成的，以响应中风的发作来解决 神经炎症和恢复动态平衡。我们的初步研究表明，NPD1在 神经细胞培养中的OGD修饰基因簇和上游电位主调节因子 神经细胞凋亡和神经炎症，改善细胞稳态，并有益于影响基因 在缺血再灌流时表达。此外，我们还表明DOC可提供神经/行为恢复， 减少脑梗塞面积，增加神经再生，促进细胞存活。的总目标是 我们的研究旨在揭示DOC在大脑中动脉阻塞中的作用机制。我们的中心假设是 DOC通过在神经元保护之前靶向选择性基因簇来促进神经元和星形胶质细胞的完整性 并受中脑星形胶质细胞源性神经营养调节的动态平衡信号整合 因子(MANF)和环指蛋白146(Idna)。具体目标1将检验DOC调控的假设 MCAO后通过调节特定基因簇促进内环境平衡和细胞存活的生物活性。我们将定义 剂量和治疗窗口，以及它们对缺血半暗带的影响，我们将定义 脂质介质对有或无组织型纤溶酶原激活剂溶栓治疗的卒中患者具有神经保护作用。 我们从我们的神经元培养数据中选择了基因，包括一些编码lncRNAs的基因，并提出 通过RT-qPCR高通量微流控工作流程确定其在有无DOC的MCAO后的表达。 特定目标2将检验MANF和IDUNA通过DOC增强丰度的假设 神经干细胞的动态平衡信号的恢复和增殖，导致神经保护。两人都支持- 针对不同神经保护机制的生存蛋白。由于NPD1的生物合成是由 我们将探讨DHA→DOC(Npd1、Rvd1和Npd1+Rvd1)→MANF与→IDUNA→的关系 通过未折叠的蛋白质反应促进生存的信号输出进行保护。将概述DOC或组合 由MANF和IDUNA驱动的未折叠蛋白质响应的输出。由于缺血性中风 UPR信号我们将LncRNAs定义为UPR的调节器和效应器，它微调UPR的输出 应激信号通路，并确定哪些特定的基因信号是MANF和/或IDUNA 依附的。拟议研究的科学前提是确定最有效的DOC或 DOC与靶向神经保护/神经修复所需基因簇的结合 脂质介体，这将为未来的临床研究提供基础的潜在干预措施，以减少 中风的直接和长期后果。

PROJECT SUMMARY / ABSTRACT Although neuroprotective strategies have shown promise, no treatment has demonstrated efficacy after stroke. This project focuses on the neuroprotective bioactivity of docosanoid (DOC) mediators: Neuroprotectin D1 (NPD1), Resolvin D1 (RvD1), and their combination (NPD1+RvD1) against ischemic and embolic experimental stroke. These lipid mediators are biosynthesized “on demand” in response to the onset of stroke to resolve neuroinflammation and restore homeostasis. Our preliminary studies show that administration of NPD1 after OGD in neuronal cell cultures modifies clusters of genes and upstream potential master regulators that decrease neuronal apoptosis and neuroinflammation, improve cell homeostasis, and that beneficially impact genes expressed in ischemia-reperfusion. In addition, we show that DOC provide neurological/behavioral recovery, reduce infarct size, increase neurogenesis, and promote cell survival after ischemic stroke. The overall goal of our studies is to uncover a mechanistic understanding of DOC action in MCAo. Our central hypothesis is that DOC foster neuronal and astrocyte integrity by targeting selective gene clusters preceding neuronal protection and by the homeostatic signaling integration regulated by the mesencephalic astrocyte-derived neurotrophic factor (MANF) and by the ring finger protein 146 (Iduna). Specific aim 1 will test the hypothesis that DOC regulate pro-homeostatic and cell survival bioactivity after MCAo by modulating specific gene clusters. We will define the doses and therapeutic window, as well as their effect on the ischemic penumbra and we will define whether the lipid mediators are neuroprotective in embolic stroke with or without tissue plasminogen activator of thrombolysis. We selected genes from our data on neuronal cultures, including, some encoding lncRNAs, and propose to define by RT-qPCR high-throughput microfluidics workflow their expression after MCAo with and without DOC. Specific aim 2 will test the hypothesis that MANF and Iduna enhanced abundance by DOC integrates homeostatic signaling restoration and proliferation of neural stem cells leading to neuroprotection. Both are pro- survival proteins that target different neuroprotective mechanisms. Since NPD1 biosynthesis is stimulated by neurotrophins, we will explore the relationship DHA→DOC (NPD1, RvD1 and NPD1+RvD1) →MANF→Iduna→ protection by unfolded protein response pro-survival signaling outputs. A DOC or combinations will outline outputs of the unfolded protein response driven by MANF and Iduna. Since ischemic stroke engages UPR signaling we will define lncRNAs as regulators and effectors of UPR that fine-tune the output of the stress signaling pathways and identify also which specific gene signatures are MANF and or Iduna dependent. The scientific premise of the proposed studies is that identification of the most effective DOC or combination of DOC to target gene clusters necessary for neuroprotection/neurorestoration modulated by the lipid mediators which will provide the basis for future clinical studies on potential interventions to reduce the immediate and long-term consequences of stroke.