Elucidating the roles of AMPK isoforms in Alzheimer's Disease Pathogenesis

项目总结/摘要 阿尔茨海默病（AD）是老年人中最常见的痴呆形式，并且可能成为一种新的 在21世纪的世纪，随着人口老龄化的迅速增长，目前没有 已知的治愈或治疗，以阻止AD的进展。此外，基本的分子机制 负责AD仍然难以捉摸。AD病理生理学中的两个关键事件是从头开始的能力受损， 蛋白质合成（mRNA翻译）和破坏细胞能量稳态。amp激活的蛋白激酶 AMPK是维持细胞能量稳态的细胞能量传感器。此外，AMPK 整合了多种信号通路（包括AKT、GSK 3、mTORC 1和eEF 2），控制从头蛋白 合成，其失调与神经退行性疾病相关的认知综合征有关 疾病包括AD。哺乳动物AMPK是一种异源三聚体蛋白，具有催化α亚基和调节α亚基， β/γ亚基。AMPK的α亚基存在两种亚型：α1和α2，它们在AD中的作用尚不清楚。通过 通过研究AD患者死后的脑组织和AD转基因小鼠模型， 发现AMPKα1的水平显著升高，而AMPKα2的水平降低。中央 当前项目的假设是AMPK同种型稳态的破坏代表了一种关键的分子机制， AD的病理生理机制。因此，本项目的目的是确定选择性AMPK是否 同种型抑制（以及随后AMPK同种型稳态的改变） 蛋白质合成和记忆形成。该项目将利用一种新的小鼠模型，其中Prkaa 1和 Prkaa 2（分别编码AMPK α1和α2亚基的基因）在兴奋性神经元中被去除， 前脑和海马发育后期，产生脑和亚型特异性条件AMPKα1 和α2基因敲除小鼠（AMPKα1 cKO和AMPKα2 cKO）。我们进一步杂交了杂合子 AMPKα1/2 cKO小鼠[AMPKα1（+/-）和AMPKα2（+/-）]与Tg 19959 AD小鼠模型（包含两个家族性 AD突变：K670 N和V717 F）产生Tg 19959/AMPKα1（+/-）和Tg 19959/AMPKα2（+/-）双 突变小鼠使用行为，电生理和生物化学方法，这里的实验将1） 阐明AMPK亚型的遗传抑制对AD相关突触可塑性损伤的影响; 2)确定AMPK同种型抑制在AD模型小鼠中学习和记忆缺陷中的作用;和 3)确定特异性AMPK同种型减少是否改善AD病理学，包括脑淀粉样蛋白 沉积和从头蛋白质合成损伤。该项目的实验结果将 有助于阐明AD病理生理学新机制， 生物标记和治疗靶点。

Project Summary/Abstract Alzheimer’s disease (AD) is the most common form of dementia in the elderly, and may become a new epidemic in the 21st century in accordance with the rapid growth of the aging population. There is currently no known cure or treatment to stop the progression of AD. Moreover, the basic molecular mechanisms responsible for AD remain elusive. Two key events in AD pathophysiology are impaired capacity of de novo protein synthesis (mRNA translation) and disrupted cellular energy homeostasis. AMP-activated protein kinase (AMPK) acts as a central cellular energy sensor to maintain cellular energy homeostasis. Furthermore AMPK integrates several signaling pathways (including AKT, GSK3, mTORC1, and eEF2) controlling de novo protein synthesis, dysregulation of which is implicated in cognitive syndromes associated with neurodegenerative diseases including AD. Mammalian AMPK is a heterotrimeric protein with a catalytic α subunit and regulatory β/γ subunit. The α subunit of AMPK exists in two isoforms: α1 and α2, and their roles in AD are unknown. By investigating brain tissue from post mortem human AD patients and a transgenic mouse model of AD, I have found that levels of AMPKα1 are dramatically increased while levels of AMPKα2 are decreased. The central hypothesis of the current project is that disruption of AMPK isoform homeostasis represents a key molecular mechanism of AD pathophysiology. Thus, the objective of this project is to determine whether selective AMPK isoform inhibition (and subsequent altering of AMPK isoform homeostasis) alleviates AD-associated deficits in protein synthesis and memory formation. This project will utilize a novel mouse model in which Prkaa1 and Prkaa2 (genes that encode AMPK α1 and α2 subunits, respectively) were removed in excitatory neurons in forebrains and hippocampus late in development, to generate brain- and isoform-specific conditional AMPKα1 and α2 knockout mice (AMPKα1 cKO and AMPKα2 cKO). We have further crossed the heterozygous AMPKα1/2 cKO mice [AMPKα1(+/-) and AMPKα2(+/-)] with Tg19959 AD mouse model (containing two familial AD mutations: K670N and V717F) to generate Tg19959/AMPKα1(+/-) and Tg19959/AMPKα2(+/-) double mutant mice. Using behavioral, electrophysiological, and biochemical methods, the experiments here will 1) elucidate the effects of genetic repression of AMPK isoforms on AD-associated synaptic plasticity impairments; 2) determine the effects of AMPK isoform suppression in learning and memory deficits in AD model mice; and 3) establish whether specific AMPK isoform reduction improves AD pathology, including brain amyloid deposition and de novo protein synthesis impairments. The experimental findings derived from this project will help elucidate a novel mechanism for AD pathophysiology, shedding a light on potentially new diagnostic biomarkers and therapeutic targets.