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Exploiting EP2 receptor biology to target seizure-related neuroinflammation selectively

利用 EP2 受体生物学选择性地靶向癫痫相关的神经炎症

基本信息

批准号：
10171930
负责人：
RAYMOND J DINGLEDINE
金额：
$ 52.02万
依托单位：
EMORY UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-06-01 至 2025-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10171930
关键词：
Ablation Address Agonist Amygdaloid structure Animal Model Anti-Inflammatory Agents Antiepileptogenic Appearance Astrocytes Autoimmune Behavioral Assay Biology Blood - brain barrier anatomy Brain Brain Injuries CX3CL1 gene Cell Survival Cells Chronic Cleaved cell Cognitive deficits Convulsants Coupled Cre driver Cyclic AMP Cyclic AMP-Dependent Protein Kinases Development Dinoprostone Disease Electroencephalography Elements Encephalitis Epilepsy Epileptogenesis Event Fractalkine Frequencies Hemin Hippocampus (Brain)Hour Human In Vitro Incidence Infection Inflammation Inflammatory Interleukin-6 Interruption Invaded Knock-out Ligands Mediating Microglia Modeling Modification Molecular Mus Myelogenous Myeloid Cells N-Methylaspartate Nerve Degeneration Neuronal Injury Neurons Open Reading Frames Oral Outcome PTGS2 gene Pathway interactions Peptides Pharmacology Pilocarpine Play Process Prosencephalon Prostaglandin E Receptor Prostaglandin Receptor Proteins Quantitative Reverse Transcriptase PCR Reaction Reporting Rodent Role Seizures Series Status Epilepticus Technology Testing Transgenes Western Blotting Work beta-arrestin butaprost cell type cyclooxygenase 2 cytokine excitotoxicity genetic approach in vivo kainate man monocyte nervous system disorder neuroinflammation neuropathology neuroprotection neurotoxicity novel prevent primary outcome receptor virtual

项目摘要

Project Summary Accumulating evidence in humans and in animal models indicates that inflammation of the brain that develops after status epilepticus (SE) may play a determinant role in long-term detrimental consequences, independent of an infection or auto-immune cause. The pathophysiological interactions among the various inflammatory molecules, and the sequence of events leading to their induction, have not yet been dissected. Previous work pointed to a role for cyclooxygenase-2 (COX-2) pathways in SE-induced inflammation, and showed that the EP2 receptor mediates much of the COX-2 effect. Our recent work suggests that PGE2 released from neurons after SE activates EP2 receptors on nearby neurons and myeloid cells, and that EP2 activation on neurons and myeloid cells might cause opposing effects. We hypothesize that EP2 activation on neurons after SE is neuroprotective, whereas EP2 activation on microglia or invading monocytes results in cytokine synthesis and subsequent development of epilepsy. Here we use a novel HaloTag technology to target neurons and myeloid cells separately with EP2 antagonists and agonists to test this hypothesis. Our specific aims are: 1. To test the hypothesis that pharmacologic block of EP2 receptors on neurons and myeloid cells has opposing effects after SE. 2. To test the hypothesis that blocking EP2 receptors on myeloid cells interferes with the process of epileptogenesis. 3. To test the hypothesis that EP2-mediated neuroprotection involves neuronal EP2 receptors, utilizes a cAMP rather than β-arrestin pathway, and requires CX3CL1 (fractalkine). To address these aims we employ in vitro culture models and in vivo SE models with novel EP2 antagonists and agonists targeted by HaloTag to neurons or microglia. Immunohistochemical, western blot, qRT-PCR, cell viability, EEG and behavioral assays are performed.

项目摘要在人类和动物模型中积累的证据表明，大脑的炎症癫痫持续状态(SE)可能在长期有害后果中起决定性作用，不受感染或自身免疫因素影响的。不同组织之间的病理生理相互作用炎症分子以及导致其诱发的一系列事件尚未被剖析。以前的工作指出了环氧合酶-2(COX-2)通路在SE诱导的炎症中的作用，以及研究表明，EP2受体在很大程度上介导了COX-2效应。我们最近的研究表明，PGE2 在SE激活附近神经元和髓细胞上的EP2受体后，神经元释放的EP2，而EP2 对神经元和髓系细胞的激活可能会导致相反的效果。我们假设EP2的激活 SE后对神经元的影响是神经保护的，而EP2对小胶质细胞或侵袭性单核细胞的激活会导致参与细胞因子的合成和癫痫的后续发展。在这里，我们使用一种新的HaloTag技术来用EP2拮抗剂和激动剂分别靶向神经元和髓系细胞来验证这一假说。我们的具体目的是：1.检验EP2受体的药物阻断对神经元和髓系细胞在SE后具有相反的作用。2.验证阻断髓系EP2受体的假说细胞干扰癫痫的发生过程。3.检验EP2介导的假说神经保护涉及神经元EP2受体，利用cAMP而不是β-arrestin途径，以及需要CX3CL1(Fractalkine)。为了达到这些目的，我们采用了体外培养模型和活体SE HaloTag靶向神经元或小胶质细胞的新型EP2拮抗剂和激动剂的模型。免疫组织化学、Western印迹、qRT-PCR、细胞活力、脑电和行为学检测。