MiR-193a-5p as a Regulator of Intersectin1-short and Clathrin-Mediated Endocytosis in Alzheimer's Disease.

MiR-193a-5p 作为阿尔茨海默病中 Intersectin1-short 和网格蛋白介导的内吞作用的调节剂。

• 批准号: 10480814
• 负责人: Sierra Smith
• 金额: $ 5.01万
• 依托单位: OREGON HEALTH & SCIENCE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-09-05 至 2023-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10480814
• 关键词: 3' Untranslated Regions; 3xTg-AD mouse; Abeta clearance; Address; Alzheimer' s Disease; Alzheimer' s disease brain; Alzheimer' s disease model; Alzheimer' s disease pathology; Alzheimer' s disease patient; Alzheimer’s disease biomarker; Amyloid beta-Protein; Astrocytes; Autopsy; Binding; Bioinformatics; Biological; Biological Assay; Biological Markers; Brain; Brain Pathology; Cause of Death; Cells; Cerebrospinal Fluid; Clathrin; Data; Dementia; Development; Diagnosis; Disease; Endocytosis; Functional disorder; Future; Gene Expression; Genetic Transcription; Genome; Hippocampus (Brain); Hormones; Human; Immunoblot Analysis; In Situ Hybridization; In Vitro; Injections; Knowledge; Lentivirus Vector; Link; Living Donors; Luciferases; Measures; Mediating; Messenger RNA; MicroRNAs; Microglia; Mus; Nerve Degeneration; Neurofibrillary Tangles; Neurologic; Neurons; Pathogenesis; Pathology; Process; Protein Isoforms; Proteins; Publishing; Regulation; Reporter; Reporting; Role; Sampling; Scaffolding Protein; Senile Plaques; Small RNA; Testing; Transferrin; Translations; United States; Wild Type Mouse; bioinformatics pipeline; brain cell; brain tissue; cell type; circulating microRNA; coated pit; cohort; differential expression; fluorescence imaging; frontal lobe; in vivo; inhibitor; interest; microRNA biomarkers; mind control; mouse model; novel therapeutic intervention; overexpression; protein expression; tool; uptake

PROJECT SUMMARY Alzheimer’s disease (AD) is the most common form of dementia and is characterized by the development of amyloid beta (Aβ) plaques and neurofibrillary tangles in the brain. Currently, the pathogenesis underlying AD is unclear and there are no reliable tools to detect early changes in the brain. Studies in the Saugstad lab revealed a specific set of microRNA (miRNA) biomarkers that are decreased in cerebrospinal fluid (CSF), and which can classify AD patients from neurologically normal controls. MiRNAs regulate post-transcriptional gene expression via complementary binding to sequences in mRNA that in turn regulate translation and expression of the resulting protein. However, there is a knowledge gap regarding which mRNAs are targeted and regulated by the AD CSF miRNAs and how these target proteins may contribute to AD pathophysiology. To address this gap, I developed a bioinformatics pipeline to predict mRNA targets of the AD CSF miRNAs, and identified Intersectin1-short (ITSN1-s) as a protein targeted by the AD CSF miRNA, miR-193a-5p. Immunoblot analysis of human post mortem hippocampus showed significant increases in ITSN1-s protein in AD relative to control. However, as ITSN1-s is expressed in multiple brain cell types, further studies that determine the specific cell type(s) with increased ITSN1-s expression in AD are needed to examine potential mechanisms underlying AD. Published studies report that ITSN-s is expressed in astrocytes and microglia, with low expression in neurons, and that ITSN1-s functions to reduce clathrin-mediated endocytosis (CME). Microglia have been shown to uptake fibrillar Aβ through a clathrin-mediated mechanism, and CME disruption in AD has been linked to reduced Aβ clearance in neurons. However, it is not known whether ITSN1-s in microglia contributes to this pathophysiology in AD. This proposal will assess the regulatory role of miR-193a-5p on ITSN1-s and its functional significance in CME and AD. In order to accomplish this, I propose the following aims: 1. establish the cellular expression of miR-193-a-5p and ITSN1-s in AD microglia and neurons; 2. investigate the interaction between miR-193a-5p and ITSN1-s mRNA and the effects on ITSN1-protein expression in vitro and in vivo; and 3. determine the role of miR-193a-5p levels on CME and Aβ uptake in AD microglia. Ultimately, these studies will provide a deeper understanding of ITSN1-s regulation in AD and how this may contribute to AD pathology. In addition, this framework can be used in future studies to evaluate the biological consequences of circulating miRNA biomarkers in AD.

项目摘要 阿尔茨海默氏病（AD）是痴呆症的最常见形式，其特征是发展 淀粉样蛋白β（Aβ）斑块和大脑中的神经原纤维缠结。目前，AD的发病机理是 不清楚，没有可靠的工具来检测大脑的早期变化。 Saugstad实验室的研究 揭示了一组特定的microRNA（miRNA）生物标志物，该标志物在脑脊液（CSF）中降低，并且 可以将AD患者分类为神经系统正常对照。 miRNA调节转录后基因 通过互补结合与mRNA中序列的互补结合，反过来调节翻译和表达 产生的蛋白质。但是，存在一个知识差距，涉及哪些mRNA针对目标，并且 受AD CSF miRNA的调节以及这些靶蛋白如何有助于AD病理生理学。到 解决这一差距，我开发了一个生物信息学管道，以预测AD CSF miRNA的mRNA靶标，并且 鉴定出由AD CSF miRNA，miR-193a-5p靶向的蛋白质互联素1-毛（ITSN1-S）。免疫印迹 对人类后海马后的分析显示，相对于AD中的ITSN1-S蛋白显着增加 控制。但是，由于ITSN1-S在多种脑细胞类型中表达，进一步的研究决定了 需要在AD中表达ITN1-S表达增加的特定细胞类型以检查潜在机制 基础广告。已发表的研究报告，ITSN-S在星形胶质细胞和小胶质细胞中表达，低 神经元中的表达，ITSN1-S的功能可减少网格蛋白介导的内吞作用（CME）。小胶质细胞 已显示通过网格蛋白介导的机制吸收原纤维Aβ，并且AD中的CME中断 我们与神经元中的Aβ清除率降低有关。但是，尚不清楚小胶质细胞中的ITSN1-S是否 在AD中有助于这种病理生理学。该提案将评估miR-193a-5p的调节作用 ITSN1-S及其在CME和AD中的功能意义。为了实现这一目标，我提出以下内容 目的：1。在AD小胶质细胞和神经元中建立miR-193-A-5p和ITSN1-S的细胞表达； 2。 研究miR-193a-5p与ITSN1-S mRNA之间的相互作用以及对ITSN1-蛋白的影响 体外和体内表达；和3。确定miR-193a-5p水平在CME和Aβ摄取AD中的作用 小胶质细胞。最终，这些研究将对AD中的ITSN1-S调节提供更深入的了解 这可能有助于AD病理。此外，该框架可以在以后的研究中用于评估 AD中循环miRNA生物标志物的生物学后果。