Single Nucleus Transcriptional Profiling of Intractable Focal Epilepsy

难治性局灶性癫痫的单核转录谱

• 批准号: 10373149
• 负责人: Peter Canoll
• 金额: $ 23.53万
• 依托单位: COLUMBIA UNIVERSITY HEALTH SCIENCES
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-01-01 至 2023-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10373149
• 关键词: Area; Astrocytes; Biological Markers; Biology; Biopsy; Brain; Cell Communication; Cell Nucleus; Cells; Characteristics; Chlorides; Cholecystokinin; Cortical Dysplasia; Data; Development; Electroencephalography; Electrophysiology (science); Epilepsy; Epileptogenesis; Frequencies; Functional disorder; Gene Expression; Gene Expression Profile; Gene Expression Profiling; Generations; Genes; Genetic Transcription; Glutamate Decarboxylase; Heterogeneity; Hippocampus (Brain); Histologic; Histopathology; Human; Image; Impairment; Individual; Interneuron function; Interneurons; Investigation; Magnetic Resonance Imaging; Microelectrodes; Microglia; Microscopic; Molecular; Molecular Target; Neurons; Operative Surgical Procedures; Partial Epilepsies; Parvalbumins; Pathologic; Pathology; Pathway interactions; Patients; Pattern; Play; Population; Process; Pyramidal Cells; Research; Resected; Resistance; Resolution; Role; Sampling; Secondary to; Seizures; Site; Testing; Therapeutic; Tissue-Specific Gene Expression; Tissues; Transcription Alteration; base; brain tissue; calbindin; cell type; differential expression; early onset; epileptic encephalopathies; improved; insight; interest; neocortical; radiological imaging; response; sodium-potassium-chloride cotransporter 1 protein; targeted treatment; transcriptome sequencing; transcriptomics

PROJECT ABSTRACT Up to 30% of individuals with epilepsy have intractable seizures. It is unclear how mechanisms underlying seizure onset and propagation operate regionally in intractable focal epilepsy, in which tissue involvement is heterogeneous and regions play different roles in seizure initiation, propagation and resistance to spread. Better understanding is needed of the biology underlying the seizure focus versus the penumbra, where excitatory discharges appear on EEG but neuronal response is limited by intact inhibition. Imbalance of inhibitory and excitatory neuronal inputs (I/E imbalance) is a primary factor underlying seizure generation and propagation, particularly interneuron dysfunction and interneuron/pyramidal cell interaction. I/E imbalance influences seizure generation in a wide array of epilepsy pathologies; investigation of I/E imbalance and interneuron function can thereby reveal pathways common to many types and causes of epilepsy. We hypothesize that cell-type specific alterations result in I/E imbalance in brain resected during surgery for intractable focal epilepsy, and that these transcriptional changes will correlate with histopathology and regional EEG. We will test this by performing single nucleus RNA sequencing (snRNAseq) on MRI-guided electrophysiologically-localized biopsies from patients undergoing epilepsy surgery. This will provide site-specific correlation of cell type-specific transcriptomic profiles with histological, radiographic, and electrophysiologic alterations. We will sample the seizure focus and penumbra to determine the differences in cellular composition and cell type-specific expression profiles, and identify alterations involved in seizure generation and propagation. Although we will focus on interneuron profiling and I/E imbalance, snRNAseq will provide data on the diversity of cell types. In AIM 1 we will use snRNAseq to identify differential expression of genes in seizure focus vs. penumbra in human brain resected during surgery for intractable focal epilepsy, focusing on interneuron vs excitatory pyramidal cell transcriptional alterations. Microelectrode recordings will provide the spatial trajectory of a seizure and define focus and penumbra. We will identify region- and cell type-specific alterations by comparing transcriptional profiles across EEG-characterized samples. We will thereby identify molecules and pathways differentially expressed in seizure focus vs. penumbra, highlighting processes likely to be primary to seizure generation. In AIM 2 we will perform immunohistochemical analyses of top target molecules identified in Aim 1, as well as molecules previously identified to play a role in I/E imbalance in epilepsy and seizure generation. Our studies will thereby provide microscopic resolution of cellular and subcellular patterns of expression within MRI-guided electrophysiologically localized biopsies. Illuminating the regional pathophysiology of seizures in intractable focal epilepsy can provide targets for therapy.

项目摘要 高达 30% 的癫痫患者患有顽固性癫痫发作。目前尚不清楚癫痫发作的机制是怎样的 顽固性局灶性癫痫的发作和传播是局部性的，其中组织受累是 异质性和区域在癫痫发作的发生、传播和抵抗蔓延方面发挥着不同的作用。更好的 需要了解癫痫发作焦点与半影的生物学基础，其中兴奋性 脑电图上出现放电，但神经元反应受到完整抑制的限制。抑制和失衡 兴奋性神经元输入（I/E 失衡）是癫痫发作产生和传播的主要因素， 特别是中间神经元功能障碍和中间神经元/锥体细胞相互作用。 I/E 不平衡影响癫痫发作 多种癫痫病理的产生； I/E 失衡和中间神经元功能的研究可以 从而揭示了许多类型和癫痫病因的共同途径。我们假设细胞类型特异性 顽固性局灶性癫痫手术期间，这些改变会导致切除大脑中的 I/E 失衡，并且这些改变 转录变化将与组织病理学和区域脑电图相关。我们将通过执行单个测试来测试这一点 对患者进行 MRI 引导的电生理局部活检的核 RNA 测序 (snRNAseq) 正在接受癫痫手术。这将提供细胞类型特异性转录组谱的位点特异性相关性 伴有组织学、放射学和电生理学改变。我们将对癫痫病灶进行采样并 半影以确定细胞组成和细胞类型特异性表达谱的差异，以及 识别癫痫发作产生和传播中涉及的改变。虽然我们将重点关注中间神经元 snRNAseq 将提供有关细胞类型多样性的数据。在 AIM 1 中我们将使用 snRNAseq 用于识别人脑切除后癫痫病灶与半暗带中基因的差异表达 在顽固性局灶性癫痫手术期间，重点关注中间神经元与兴奋性锥体细胞转录 变更。微电极记录将提供癫痫发作的空间轨迹并定义焦点和 半影。我们将通过比较跨区域的转录谱来识别区域和细胞类型特异性的改变 脑电图特征样本。因此，我们将识别癫痫发作中差异表达的分子和途径 焦点与半暗带，突出显示可能是癫痫发作产生的主要过程。在 AIM 2 中我们将执行 对目标 1 中确定的顶级靶分子以及之前的分子进行免疫组织化学分析 确定在癫痫和癫痫发作的 I/E 失衡中发挥作用。我们的研究将因此提供 MRI 引导的电生理学中细胞和亚细胞表达模式的微观分辨率 局部活检。阐明难治性局灶性癫痫发作的区域病理生理学可以提供 治疗的目标。