Studies of Ahnak pathways in endothelial cells and blood-brain barrier regulation

内皮细胞 Ahnak 通路和血脑屏障调节的研究

基本信息

批准号：
10866696
负责人：
Yong Kim
金额：
$ 26.69万
依托单位：
RUTGERS BIOMEDICAL AND HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-03-25 至 2025-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10866696
关键词：
ANXA2 gene Affect Affinity Chromatography Antidepressive Agents Astrocytes Biochemical Biological Assay Blood Blood - brain barrier anatomy Blood Cells Blood Vessels Brain region CRISPR/Cas technology Calcium Channel Candidate Disease Gene Cells Central Nervous System Cerebrovascular system Communication Contrast Media Data Dyes Endothelial Cells Enterobacteria phage P1 Cre recombinase Exclusion Functional disorder Gadolinium Genes Hippocampus Homeostasis Horseradish Peroxidase Immunohistochemistry Knock-in Knock-out Knockout Mice LoxP-flanked allele MRI Scans Measures Mediating Mental disorders Messenger RNA Methods Molecular Mood Disorders Moods Mus Neurons Ontology Outcome Outcome Study Pathway interactions Pattern Penetration Pericytes Psychiatry Regulation Ribosomes Role Silicon Silicones Tight Junctions Translating Transmission Electron Microscopy Vascular System Visualization Wild Type Mouse behavioral phenotyping blood-brain barrier function blood-brain barrier permeabilization brain parenchyma brain tissue density dentate gyrus depressive behavior differential expression granule cell insight intravenous administration intravenous injection meter neural circuit neuronal circuitry novel novel strategies prevent protein complex public health relevance transcriptome sequencing voltage

项目摘要

Modified Project Summary/Abstract Section Endothelial cells (ECs) in blood vessels within brain parenchyma tightly regulate the function of the blood-brain barrier (BBB) and the communication between the vascular system and neighboring neurons to maintain circuitry homeostasis. In a recent study, we elucidated that Ahnak is a novel regulator of depressive behavior and is an endogenous neuronal scaffolder of the S100A10 (p11)/Anxa2 protein complex and L-type voltage-gated calcium channels, both of which have been implicated in the pathophysiology of psychiatric disorders. In addition to neuronal expression, Ahnak is highly expressed in ECs in blood vessels in the brain. However, Ahnak’s role in the ECs in the regulation of BBB function has not yet been investigated. Our preliminary data indicate that EC-specific Ahnak knockout (KO) mice display an antidepressant-like behavioral phenotype. Because ECs are responsible for BBB permeability, we hypothesize that Ahnak deletion and alterations of Ahnak downstream pathways in ECs modulate BBB permeability and thereby affect neuronal circuit activities. We aim to elucidate Ahnak-mediated molecular and functional pathways in the ECs. We also aim to investigate the roles of Ahnak in ECs in the regulation of BBB function and the communication between the vascular system and neural circuits in the ventral hippocampus. First, we will investigate Ahnak downstream genes and pathways in ECs using an EC-selective TRAP (Translating Ribosome Affinity Purification)/RNA-seq approach. By investigating differentially expressed genes in EC-specific Ahnak KO mice (floxed Ahnak mice crossed with Tek-Cre mice) compared to wild type (WT) controls, we aim to identify downstream molecules and Ahnak-mediated functional pathways (Specific Aim 1). Second, we will investigate the effect of EC-specific Ahnak KO on the BBB permeability in the ventral hippocampus using magnetic resonance imaging (MRI) scans, immunohistochemistry, biochemical assays and transmission electron microscopy (Specific Aim 2). Lastly, we will investigate the effect of Ahnak deletion in ECs on neuronal activity in the ventral dentate gyrus (vDG) using high-density silicon probes recordings of WT and EC-in-vDG (EC[vDG])-specific Ahnak KO mice. EC[vDG]-specific Ahnak deletion will be achieved by a CRISPR/Cas9 approach (Specific Aim 3). The outcome of the proposed studies will identify novel molecular factors and mechanisms regulating homeostasis of the ventral hippocampal circuitry.

修改后的项目摘要/摘要部分脑实质内血管中的内皮细胞（EC）严格调节血脑屏障（BBB）的功能以及血管系统与邻近神经元之间的通讯，以维持电路稳态。是一种新型的抑郁行为调节剂，是 S100A10 (p11)/Anxa2 蛋白复合物和 L 型电压门控钙通道的内源性神经元支架，两者均是除了神经元表达之外，Ahnak 在大脑血管中的 EC 中也高度表达，但 Ahnak 在调节 BBB 功能中的作用尚未得到研究。初步数据表明，EC 特异性 Ahnak 敲除 (KO) 小鼠表现出抗抑郁样行为表型，因为 EC 负责 BBB 通透性，我们认为 Ahnak 缺失和改变。 EC 中 Ahnak 下游通路调节 BBB 通透性，从而影响神经回路活动我们的目的是阐明 EC 中 Ahnak 介导的分子和功能通路。首先，我们将使用 EC 选择性 TRAP 研究 EC 中的 Ahnak 下游基因和通路。（翻译核糖体亲和纯化）/RNA-seq 方法通过研究 EC 特异性 Ahnak KO 小鼠（与 Tek-Cre 小鼠杂交的 floxed Ahnak 小鼠）与野生型 (WT) 对照相比的差异表达基因，我们的目标是识别下游分子。和 Ahnak 介导的功能通路（具体目标 1），我们将利用磁力研究 EC 特异性 Ahnak KO 对腹侧海马 BBB 通透性的影响。最后，我们将使用高密度硅探针研究 EC 中 Ahnak 缺失对腹侧齿状回 (vDG) 神经元活动的影响。 WT 和 EC-in-vDG (EC[vDG]) 特异性 Ahnak KO 小鼠的记录。删除将通过 CRISPR/Cas9 方法实现（具体目标 3）。拟议研究的结果将确定调节腹侧海马回路稳态的新分子因子和机制。