Calcineurin and inflammatory signaling processes in aging and Alzheimer's Disease

衰老和阿尔茨海默病中的钙调神经磷酸酶和炎症信号传导过程

基本信息

批准号：
8657965
负责人：
Christopher Mark Norris
金额：
$ 29.96万
依托单位：
UNIVERSITY OF KENTUCKY
依托单位国家：
美国
项目类别：
财政年份：
2006
资助国家：
美国
起止时间：
2006-09-01 至 2017-03-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8657965
关键词：
APP-PS1 Age Aging Alzheimer&apos s Disease Alzheimer&apos s disease model Animals Astrocytes Automobile Driving Avoidance Learning Behavior Behavioral Biochemical Process Biological Assay Biological Markers Brain Brain Diseases Calcineurin Cells Ceramics Cognitive Data Dependence Dependovirus Deposition Development Disease Electrodes Electrophysiology (science)Enzyme-Linked Immunosorbent Assay Enzymes Funding Gene Delivery Glial Fibrillary Acidic Protein Glutamate Receptor Glutamate Transporter Glutamates Hippocampus (Brain)Human Hypertrophy Impaired cognition Implant In Vitro Inflammatory Ipsilateral Label Measurement Measures Methods Microelectrodes Microglia Mus NFAT Pathway Nerve Degeneration Nervous System Physiology Neurodegenerative Disorders Neurologic Dysfunctions Outcome Measure Pathologic Pathology Pathway interactions Phenotype Process Production Protein phosphatase Proteins Reaction Regulation Resolution Rodent Model Role Signal Transduction Slice Staging Synapses Synaptic plasticity Techniques Testing Therapeutic Time Tissues Transgenic Mice Transgenic Organisms Western Blotting Wild Type Mouse adeno-associated viral vector aging brain amyloid pathology astrogliosis base beta-site APP cleaving enzyme 1 cytokine functional outcomes improved in vivo inhibitor/antagonist innovation interest mouse model neuroinflammation neuropathology neuroprotection novel strategies prevent promoter protein expression relating to nervous system research study response signal processing synaptic function transcription factor treatment strategy vector

项目摘要

DESCRIPTION (provided by applicant): The experiments in this project use innovative gene delivery techniques and a multipronged approach to assess the fundamental role of astrocyte activation in neurologic function in an intact mouse model of Alzheimer's disease (AD). Studies use adeno-associated virus vectors (AAV) bearing the astrocyte-specific promoter Gfa2 to target the protein phosphatase calcineurin (CN) and NFAT transcription factors in astrocytes of wild-type and APP/PS1 mice. AAV-Gfa2 vectors are bilaterally delivered to the hippocampus at different ages/disease stages and mice are assessed on different AD biomarkers. In Aim 1, cognitive status is assessed using the active avoidance task, while synaptic function is evaluated using hippocampal slice electrophysiology and Western blot measures of synaptic proteins. In Aim 2, hippocampal glutamate regulation is investigated using ceramic enzyme-based microelectrode arrays and measures of glutamate transporter levels. In Aim 3, levels of glial activation and neuroinflammation are determined with immunohistochemical (IHC) analyses and assessment of cytokine levels using Multiplex ELISAs. In Aim 4, IHC is used to determine the extent of A? deposition, while ELISAs are used to quantify levels of A?40 and A?42 in soluble and insoluble hippocampal tissue fractions, and Westerns used to assess BACE protein expression. AAV-Gfa2 vectors encode either potent inhibitors or activators of CN/NFAT signaling and therefore will determine the necessity and sufficiency of this astrocytic pathway in driving and/or maintaining neurologic dysfunction in AD mice. These studies provide a highly novel approach to the study of activated astrocytes and could have a major impact on the development of treatment strategies for AD and other neurodegenerative conditions.

描述（由申请人提供）：该项目的实验使用创新的基因传递技术和多管齐下的方法来评估星形胶质细胞激活在完整的阿尔茨海默病（AD）小鼠模型的神经功能中的基本作用。研究使用带有星形胶质细胞特异性启动子 Gfa2 的腺相关病毒载体 (AAV) 来靶向野生型和 APP/PS1 小鼠星形胶质细胞中的蛋白磷酸酶钙调磷酸酶 (CN) 和 NFAT 转录因子。 AAV-Gfa2 载体在不同年龄/疾病阶段被双侧递送至海马，并对小鼠进行不同 AD 生物标志物的评估。在目标 1 中，使用主动回避任务评估认知状态，同时使用海马切片电生理学和突触蛋白的蛋白质印迹测量评估突触功能。在目标 2 中，使用基于陶瓷酶的微电极阵列和谷氨酸转运蛋白水平的测量来研究海马谷氨酸调节。在目标 3 中，通过免疫组织化学 (IHC) 分析和使用多重 ELISA 评估细胞因子水平来确定神经胶质活化和神经炎症的水平。在目标 4 中，IHC 用于确定 A？沉积，而 ELISA 用于定量可溶性和不溶性海马组织部分中 Aβ40 和 Aβ42 的水平，Western 用于评估 BACE 蛋白表达。 AAV-Gfa2 载体编码 CN/NFAT 信号传导的有效抑制剂或激活剂，因此将确定该星形胶质细胞途径在驱动和/或维持 AD 小鼠神经功能障碍方面的必要性和充分性。这些研究为研究活化星形胶质细胞提供了一种高度新颖的方法，可能对 AD 和其他神经退行性疾病治疗策略的制定产生重大影响。