Deciphering Mechanisms of Astrocyte-BBB Interaction in Normal and Ischemic Stroke

解读正常和缺血性中风中星形胶质细胞-BBB相互作用的机制

基本信息

批准号：
10585849
负责人：
Hyun Kyoung Lee
金额：
$ 45.68万
依托单位：
BAYLOR COLLEGE OF MEDICINE
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-01-01 至 2027-12-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10585849
关键词：
Ablation Address Adult Anatomy Area Arginine Astrocytes Blood - brain barrier anatomy Blood Vessels Brain Brain Injuries Brain region CCL2 gene Cause of Death Cell Communication Cell physiology Cells Cerebral Ischemia Conditioned Culture Media Coupled Data Degenerative Disorder Disease Endothelial Cells Endothelium Exhibits Extravasation Genetic Gliosis Goals Health Homeostasis Impairment In Vitro Infarction Intercellular Junctions Ischemic Stroke Knowledge Maintenance Malignant Neoplasms Mediating Metabolic Metabolism Molecular Morphology Mus Neurodevelopmental Disorder Neuroglia Oxides Paracrine Communication Pathogenesis Pathologic Pathway interactions Patients Physiological Physiology Play Production Property Role Signal Pathway Signal Transduction Sodium-Bicarbonate Symporters Stroke Testing Time Transgenic Mice Variant awake blood-brain barrier disruption blood-brain barrier function cell type central nervous system injury chemokine cytokine extracellular gain of function in vivo loss of function motor disorder mouse model multiple omics nervous system disorder novel pH Homeostasis post stroke prevent response stroke patient stroke risk

项目摘要

SUMMARY AND ABSTRACT Astrocytes are the most diverse glial cell type and maintain essential interactions with endothelial cells to form the blood-brain barrier (BBB). The BBB plays a major role in CNS homeostatic function, while dysregulation of the astrocytic BBB contributes to a spectrum of neurological disorders, ranging from neurodevelopmental and degenerative diseases to CNS injury and malignancy. Particularly, pathologic astrocyte-BBB interactions contribute to ischemic stroke, which is the 5th leading cause of death in the U.S. However, the mechanisms underlying maintenance of BBB integrity, both in health and in diseases such as stroke, remain poorly defined. The overarching goal of this proposal is to elucidate novel targets and associated signaling pathways influencing normal astrocyte-BBB function and to leverage this knowledge to define key mechanisms for BBB disruption after ischemic cerebral stroke. This proposal focuses on an astrocyte-enriched sodium-bicarbonate cotransporter 1, Slc4a4, which was previously identified as a glial-specific regulator of both intracellular and extracellular pH. While pH homeostasis is essential for brain function and patients carrying Slc4a4 variants can suffer ischemic stroke, a regulatory role of Slc4a4 in astrocyte-BBB integrity remains unknown. To begin addressing this knowledge gap, we generated new transgenic mouse lines that temporally ablate Slc4a4 in astrocytes. Using this genetic mouse model, we showed that loss of Slc4a4 significantly reduces astrocytic morphological complexity and generates enlarged blood vessels with disrupted endothelial junctions. Our profiling analyses of astrocytic Slc4a4-deficient mice and conditioned media of Slc4a4 ablated astrocytes revealed increased CCL2 production and secretion coupled with dysregulated arginine-nitro oxide (NO) metabolism, further supporting a crucial role for Slc4a4 in astrocyte-BBB integrity. Using an ischemic stroke mouse model, we found loss of Slc4a4 exacerbates stroke-induced motor dysfunction and increases infarct area coupled with impaired reactive gliosis which results in BBB disruption, which is rescued by inhibition of CCL2 in vivo. Based on these preliminary data, our central hypothesis is that Slc4a4 functions to maintain astrocyte-BBB interactions and prevent progressive BBB leakage in ischemic stroke, in part by inhibiting the chemokine CCL2. To address our hypothesis, we will first determine how Slc4a4 regulates astrocyte morphology and physiology in the adult brain (Aim 1). Second, we will determine the role of CCL2 in Slc4a4-dependent BBB maintenance in the adult brain (Aim 2). Upon completion, these studies will establish whether and how Slc4a4-deficient astrocytes influence anatomical and physiological properties of astrocytes, and will elucidate how Slc4a4 regulates astrocytic modulation of endothelial cell and BBB integrity via NO metabolism that drive astrocytic CCL2 signaling in the adult brain. Lastly, we will define the Slc4a4 pathway in the maintenance of BBB integrity following ischemic stroke (Aim 3). Together, elucidating the Slc4a4 pathway could reveal fundamental mechanisms controlling astrocyte morphology, physiology, and secreted molecules, and it will advance our understanding of astrocyte-BBB interactions after stroke.

摘要和摘要星形胶质细胞是最多样化的神经胶质细胞类型，并保持与内皮细胞的基本相互作用以形成血脑屏障（BBB）。 BBB在CNS稳态功能中起主要作用，而失调星形胶质细胞BBB有助于神经系统疾病，从神经发育和 CNS损伤和恶性肿瘤的退化性疾病。特别是病理星形胶质细胞BBB相互作用有助于缺血性中风，这是美国第五大死亡原因，但是，这些机制在健康和中风等疾病中，BBB完整性的基本维护仍然很差。该提案的总体目标是阐明新的目标和相关的信号通路正常的星形胶质细胞BBB功能，并利用这些知识来定义BBB中断的关键机制缺血性脑卒中。该提案重点是富含星形胶质细胞的钠双碳酸盐氧化钠 1，SLC4A4，以前被确定为细胞内和细胞外pH的神经胶质特异性调节剂。虽然pH稳态对于大脑功能至关重要，而携带SLC4A4变体的患者可能会缺血中风，SLC4A4在星形胶质细胞BBB完整性中的调节作用仍然未知。开始解决这个问题知识差距，我们生成了新的转基因小鼠线，这些小鼠系在星形胶质细胞中暂时烧开SLC4A4。使用此遗传小鼠模型，我们表明SLC4A4的损失显着降低了星形胶质形态的复杂性并产生肿大的血管，内皮连接处干扰。我们对星形胶质细胞的分析分析 SLC4A4缺陷小鼠和SLC4A4烧蚀的星形胶质细胞的条件介质显示CCL2产生增加并分泌与失调的精氨酸氧化物（NO）代谢相结合，进一步支持至关重要的作用对于星形胶质细胞BBB完整性中的SLC4A4。使用缺血性卒中小鼠模型，我们发现SLC4A4的损失中风诱导的运动功能障碍并增加梗塞区域以及反应性神经胶质的受损，从而导致在BBB中断中，通过抑制CCL2在体内可以挽救。基于这些初步数据，我们的中央假设是SLC4A4的功能可维持星形胶质细胞BBB相互作用并防止进行性BBB泄漏在缺血性中风中，部分通过抑制趋化因子CCL2。为了解决我们的假设，我们将首先确定 SLC4A4如何调节成年大脑中的星形胶质细胞形态和生理学（AIM 1）。第二，我们将确定 CCL2在成年大脑中依赖SLC4A4依赖性BBB维持中的作用（AIM 2）。完成后，这些研究将确定SLC4A4缺陷的星形胶质细胞是否以及如何影响解剖学和生理星形胶质细胞的特性，并将阐明SLC4A4如何调节内皮细胞的星形细胞调节和 BBB完整性通过无代谢，驱动成人大脑中星形胶质细胞CCL2信号传导。最后，我们将定义缺血性中风后BBB完整性维持中的SLC4A4途径（AIM 3）。一起阐明 SLC4A4途径可以揭示控制星形细胞形态，生理学和分泌的分子，它将提高我们对中风后星形胶质细胞BBB相互作用的理解。