Differentially expressed hippocampal miRNAs with age and cognitive decline

描述(由申请人提供)：随着年龄的增长，认知能力下降会降低老年人的独立性、生活质量和生产力。虽然比阿尔茨海默氏症和帕金森氏症等神经退行性疾病更常见，但认知能力下降的原因仍不清楚。这项研究的中心假设是miRNAs的差异表达随着年龄和认知能力的下降而对突触的可塑性和认知产生负面影响，这是通过调节蛋白质表达来实现的。我们之前的研究已经利用Morris水迷宫测试对Fischer 344 x Brown挪威(F1)大鼠进行认知分层，确定了随着年龄和认知能力的下降，海马区mRNA和蛋白质表达的变化。有趣的是，在比较成年(12个月)和老年(26个月)认知正常的大鼠和老年认知受损的大鼠时，我们发现了一些与神经元可塑性相关的蛋白质表达的变化，这些变化不是在转录水平上调节的。这表明了包括miRNAs在内的蛋白质表达调控的替代机制的作用。在使用相同模型和行为测试的先导研究中，我们发现了随年龄调节的miRNAs，以及认知正常和受损的老年动物之间特定调节的miRNAs。这些miRNAs的预测靶标包括许多我们已经确定为下调但其转录水平没有变化的蛋白质。这表明miRNAs作为海马区蛋白表达的调节机制之一，随着认知能力的下降和衰老而改变。为了扩展我们的初步结果，在第一个特定目标中，将测定所有已知的大鼠miRNAs在成年和老年认知分层Fischer 344 x Brown挪威大鼠的CA1、CA2/3和DG海马亚区的表达。这将首次对衰老和认知障碍的海马区miRNA表达进行全基因组检查。差异表达的miRNAs将通过定量聚合酶链式反应确认，潜在的靶点将通过电子分析确定。在目标预测之后，所有的miRNA表达和目标数据将与我们现有的关于衰老和认知衰退的mRNA和蛋白质表达数据集整合，以确定特定miRNAs可能的调控点。第二个特定目标将确定由差异表达的miRNAs调控的特定蛋白质集。在神经元细胞培养中，受认知能力下降或衰老调节的特定miRNAs将过度/不足表达，以评估特定和整体蛋白质表达的变化。直接/间接调控的区别将通过使用被调控基因的3‘UTR3’UTR的报告分析来确定。了解与衰老和认知功能下降有关的miRNAs，以及它们确定的调控靶点，将有助于理解miRNA表达变化对可塑性和认知的潜在结果。 公共卫生相关性：随着年龄的增长，认知能力下降会降低老年人的独立性、生活质量和生产力。尽管人们对阿尔茨海默氏症和帕金森氏症等神经退行性疾病的机制知之甚少，但对导致认知能力随年龄增长而下降的分子变化却知之甚少。这个项目将建立在之前对海马体中与年龄相关的认知下降的突触机制的研究基础上。本文提出的研究将检验一种假设，即miRNA的表达随着年龄的增长和/或认知能力的下降而变化，调节突触蛋白表达的变化，从而导致神经传递效率下降。这项研究将有助于阐明认知衰退的原因，并有可能为新疗法的开发提供目标，这些疗法将增加老年人的认知健康跨度。

DESCRIPTION (provided by applicant): Cognitive decline with aging reduces independence, quality of life, and productivity in older individuals. While much more common than neurodegenerative diseases, such as Alzheimer's and Parkinson's, the etiology of cognitive decline remains unknown. The central hypothesis of this proposed study is that differential expression of miRNAs with age and cognitive decline negatively impacts synaptic plasticity and cognition through the regulation of protein expression. Our previous studies have identified hippocampal mRNA and protein expression changes with aging and cognitive decline utilizing Fischer 344 x Brown Norway (F1) rats cognitively stratified by Morris Water Maze testing. Interestingly, in comparisons of Adult (12 month) and Aged (26 Month) Cognitively Intact, and Aged Cognitively Impaired rats we have identified a number of changes in protein expression related to neuronal plasticity that are not regulated at the transcript level. This suggests a role for alternate mechanisms of protein expression regulation, including miRNAs. In pilot studies using the same model and behavioral testing we have identified miRNAs regulated with age and miRNAs specifically regulated between cognitively intact and impaired aged animals. The predicted targets of these miRNAs included many of those proteins that we have identified as down regulated but whose transcript levels are unchanged. This suggests a role for miRNAs as one of the regulatory mechanisms underlying the hippocampal protein expression changes with cognitive decline and aging. To expand on our preliminary results, in the first specific aim, hippocampal expression of all known rat miRNAs will be determined in CA1, CA2/3 and DG hippocampal sub regions of Adult and Aged cognitively stratified Fischer 344 x Brown Norway rats. This will provide the first genome-wide examination of hippocampal miRNA expression with aging and cognitive impairment. Differentially expressed miRNAs will be confirmed by qPCR and potential targets will be determined by in silico analysis. Following target prediction, all miRNA expression and target data will be integrated with our existing mRNA and protein expression datasets on aging and cognitive decline to identify likely points of regulation by specific miRNAs. The second specific aim will determine specific sets of proteins regulated by differentially expressed miRNAs. Specific miRNAs regulated with cognitive decline or aging will be over/under-expressed in neuron cell culture to assess both specific and global changes in protein expression. Differentiation of direct/indirect regulation will be determined through reporter assays using the 3' UTR of regulated genes. Knowing not only the miRNAs regulated with aging and cognitive decline but their confirmed regulatory targets will allow for an understanding of the potential outcome of altered miRNA expression on plasticity and cognition. PUBLIC HEALTH RELEVANCE: Cognitive decline with aging reduces independence, quality of life, and productivity in older individuals. Although much is known about the mechanisms underlying neurodegenerative diseases such as Alzheimer's and Parkinson's, very little is known concerning the molecular changes that contribute to cognitive decline with age. This project will build upon previous research of synaptic mechanisms of age-associated cognitive decline in the hippocampus. The studies proposed here will examine the hypothesis that miRNA expression changes with aging and/or cognitive decline regulate changes in synaptic protein expression, resulting in decreased neurotransmission efficiency. This research will help elucidate the etiology of cognitive decline and potentially provide targets for the development of new therapies which will increase the cognitive health span for older individuals.