The STAT3 response of excitatory neurons to epileptogenic brain injury

兴奋性神经元对致癫痫性脑损伤的 STAT3 反应

• 批准号: 10467510
• 负责人: Amy R. Brooks-Kayal
• 金额: $ 67.14万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-05-01 至 2027-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10467510
• 关键词: Acute; Adverse effects; Animal Model; Astrocytes; Attenuated; Binding Sites; Brain; Brain Diseases; Brain Injuries; Brain-Derived Neurotrophic Factor; Ca(2+)-Calmodulin Dependent Protein Kinase; Cell Nucleus; Cells; Chromatin; Computer Models; Cyclic AMP; DNA; Data; Development; Disease; Disease Progression; Down-Regulation; Electrophysiology (science); Encephalitis; Enterobacteria phage P1 Cre recombinase; Epilepsy; Epileptogenesis; Frequencies; GABA-A Receptor; GRM5 gene; Gene Expression Regulation; Gene Silencing; Genes; Genetic Transcription; Genome; Genomics; Glutamate Receptor; Glutamates; Hippocampus (Brain); Human; Immunity; Impaired cognition; Individual; Inflammation; Inflammatory; Injections; Intervention; JAK2 gene; Janus kinase; Kainic Acid; Knock-out; Knowledge; Laboratories; Long-Term Depression; Malignant neoplasm of brain; Mediating; Mediator of activation protein; Medical; Memory impairment; Metabotropic Glutamate Receptors; Microglia; Modeling; Molecular; Monitor; Morphology; Mus; N-Methyl-D-Aspartate Receptors; Nerve Degeneration; Neuroglia; Neuronal Plasticity; Neurons; Neuropharmacology; Pathogenesis; Pathologic; Pathway interactions; Patients; Pharmacology; Pilocarpine; Population; Predisposition; Property; Prosencephalon; Protein Tyrosine Kinase; RNA; Receptor Signaling; Recurrence; Reporting; Role; STAT protein; STAT3 gene; Seizures; Signal Transduction; Slice; Status Epilepticus; Symptoms; Synaptic plasticity; Tamoxifen; Temporal Lobe Epilepsy; Testing; Therapeutic Intervention; Tissues; Transgenic Organisms; Wild Type Mouse; biocytin; brain cell; calmodulin-dependent protein kinase II; cell type; chromatin remodeling; conditioned fear; dimer; excitatory neuron; gene network; gene repression; glial activation; granule cell; inflammatory marker; inhibitor; inhibitory neuron; kainate; molecular imaging; mouse model; multiple omics; neural circuit; neuroinflammation; neuronal excitability; neurotransmission; new therapeutic target; prevent; promoter; receptor; relating to nervous system; response; response to injury; synaptogenesis; targeted treatment; transcription factor; transcriptome; transcriptome sequencing; transcriptomics

Abstract Temporal lobe epilepsy (TLE) is a progressive disorder mediated by pathological changes in molecular cascades and neural circuit remodeling in the hippocampus resulting in increased susceptibility to spontaneous seizures and cognitive dysfunction. Targeting these cascades could prevent or reverse symptom progression and has the potential to provide viable disease-modifying treatments that could reduce the portion of TLE patients (>30%) not responsive to current medical therapies. The Janus Kinase/Signal Transducer and Activator of Transcription (JAK/STAT) pathway has recently been implicated in the pathogenesis of TLE. This pathway is known to be involved in inflammation and immunity, and to be critical for neuronal functions such as synaptic plasticity and synaptogenesis. Our laboratories previously showed that a STAT3 inhibitor, WP1066, could greatly reduce the number of spontaneous recurrent seizures (SRS) in an animal model of pilocarpine-induced status epilepticus (SE). While this suggests promise for JAK/STAT inhibitors as disease-modifying therapies, the potential adverse effects of systemic or global CNS pathway inhibition limits their use. Development of more targeted therapeutics will require a detailed understanding of JAK/STAT-induced epileptogenic responses in different cell types. To this end, we have developed a new transgenic line where dimer-dependent STAT3 signaling is functionally knocked out (fKO) by tamoxifen-induced Cre expression specifically in forebrain excitatory neurons (eNs) via the Calcium/Calmodulin Dependent Protein Kinase II alpha (CamK2a) promoter. We now report that STAT3 KO in excitatory neurons (eNSTAT3fKO) markedly reduces the progression of epilepsy (SRS frequency) in the intrahippocampal kainate (IHKA) TLE model and protects mice from kainic acid (KA)-induced memory deficits as assessed by Contextual Fear Conditioning. Using data from bulk hippocampal tissue RNA-sequencing, we further discovered a transcriptomic signature for the IHKA model that contains a substantial number of genes, particularly in synaptic plasticity and inflammatory gene networks, that are down-regulated after KA-induced SE in wild-type but not eNSTAT3fKO mice. In this application, we will test the hypothesis that STAT3 signaling in excitatory neurons is a key driver of epilepsy progression via the selective silencing of genes that regulate synaptic plasticity and neuroinflammation. With an integration of open discovery using multiomics and quantitative molecular imaging (Aims 1 and 3), in combination with electrophysiology and neuropharmacology (Aim 2), we will elucidate the genome’s response to injury (24 h and 4 wks after IHKA) within different cell types and determine why STAT3 KO in eNs inhibits disease progression after KA injection by identifying direct and indirect effects of loss of eNSTAT3 expression on both excitatory and inhibitory neurons. We will also determine the relationship between eNSTAT3 signaling and glial activation by examining effects of eNSTAT3KO on the glial transcriptome and inflammatory markers of microglia and astrocytes. Our results will ascertain if cell-type specific modulation of STAT3 signaling or its downstream targets are promising strategies for therapeutic intervention.

抽象的 颞叶癫痫（TLE）是一种由分子级联病理变化介导的进行性疾病 海马体的神经回路重塑导致自发性癫痫发作的易感性增加 和认知功能障碍。针对这些级联反应可以预防或逆转症状进展，并具有 提供可行的疾病缓解治疗的潜力，可以减少 TLE 患者的比例 (>30%) 对当前的医学疗法没有反应。 Janus 激酶/信号转导器和转录激活器 (JAK/STAT) 通路最近被认为与 TLE 的发病机制有关。已知该途径是 参与炎症和免疫，并对神经元功能（例如突触可塑性和 突触发生。我们的实验室之前表明，STAT3 抑制剂 WP1066 可以大大降低 毛果芸香碱诱发的癫痫持续状态动物模型中自发性复发性癫痫发作 (SRS) 的次数 （SE）。虽然这表明 JAK/STAT 抑制剂有望作为疾病缓解疗法，但潜在的不利影响 全身或整体中枢神经系统通路抑制的影响限制了它们的使用。开发更有针对性的治疗方法 需要详细了解不同细胞类型中 JAK/STAT 诱导的致癫痫反应。到 为此，我们开发了一种新的转基因品系，其中二聚体依赖性 STAT3 信号传导在功能上发挥作用 被他莫昔芬诱导的 Cre 表达敲除（fKO），特别是在前脑兴奋性神经元（eNs）中 钙/钙调蛋白依赖性蛋白激酶 II α (CamK2a) 启动子。我们现在报告 STAT3 KO 兴奋性神经元 (eNSTAT3fKO) 显着降低癫痫的进展（SRS 频率） 海马内红藻氨酸 (IHKA) TLE 模型并保护小鼠免受红藻氨酸 (KA) 诱导的记忆缺陷 通过情境恐惧调节进行评估。使用大量海马组织 RNA 测序数据，我们 进一步发现了 IHKA 模型的转录组特征，其中包含大量基因， 特别是在突触可塑性和炎症基因网络中，这些网络在 KA 诱导 SE 后下调 在野生型小鼠中，但在 eNSTAT3fKO 小鼠中则不然。在此应用中，我们将测试 STAT3 信号传导的假设 兴奋性神经元是癫痫进展的关键驱动因素，通过选择性沉默调节基因 突触可塑性和神经炎症。通过使用多组学和开放发现的集成 定量分子成像（目标 1 和 3），结合电生理学和神经药理学 （目标 2），我们将阐明不同细胞类型中基因组对损伤（IHKA 后 24 小时和 4 周）的反应 并通过鉴定直接和 eNSTAT3 表达缺失对兴奋性和抑制性神经元的间接影响。我们还将确定 通过检查 eNSTAT3KO 对神经胶质细胞的影响，研究 eNSTAT3 信号传导与神经胶质细胞激活之间的关系 小胶质细胞和星形胶质细胞的转录组和炎症标记物。我们的结果将确定细胞类型是否具有特异性 STAT3 信号传导或其下游靶标的调节是有前途的治疗干预策略。