Organellar pH shifts in Alzheimer's disease from simulation & experiment

描述（由申请人提供）：目前尚不清楚在阿尔茨海默病（AD）中，淀粉样蛋白-ß肽（a ßx）主要在脑神经细胞（哪些细胞器）中产生，以及a ßx的过量产生是否由于淀粉样蛋白前体蛋白（APP）和/或在AD中处理APP的ß-分泌酶的错误转运/错误转运。在AD中，存在脑酸中毒（例如pH 6.6）和细胞内pH值降低。由于ß-secretase具有最佳酸性pH值，因此确定细胞内酸中毒是否也导致处理APP的细胞器pH值降低是很重要的。提出的工作的目的是：(1)通过计算机模拟粗细胞器模型（脂质体）的管腔pH变化如何影响a ß42自聚集和a ß42-脂质相互作用，获得分子水平的理解；(2)建立早期和晚期内核体（LE）、溶酶体（LY）、&高尔基体(的细胞器ßx可能产生)在胚胎和成年小鼠大脑神经元的函数(一)每个细胞器-类型的靠近身体细胞,(3)描述这些分布如何变化在转基因小鼠模型(5 xfad)的家族性阿尔茨海默氏症(时尚)的函数(b)老鼠的年龄和(c)总值脑区(海马& isocortex)相比,控制,和(4)描述colocalization如何应用,ßx和

DESCRIPTION (provided by applicant): It is unclear where within brain neurons (which organelles) the amyloid-ß peptide (Aßx) is predominantly produced in Alzheimer's disease (AD) & whether or not the overproduction of Aßx that occurs is due to a mistrafficking/misrouting of the amyloid precursor protein (APP) &/or the ß-secretase enzyme that processes APP in AD. In AD, there is cerebral acidosis (e.g., pH 6.6) & a decreased intracellular pH.1 Since ß-secretase has an acidic pH optimum, it is important to determine if the intracellular acidosis also leads to decrease in the pH of organelles within which APP is processed. The Aims of the proposed work are to (1) gain a molecular level understanding from computer simulations of how changes in the lumenal pH of crude organelle models (liposomes) impact the Aß42 self-aggregation & Aß42-lipid interactions, to (2) establish the baseline pH distribution of the early & late endosome (LE), lysosome (LY), & Golgi apparatus (the organelles within which Aßx is potentially produced) within embryonic & adult mouse brain neurons as a function of (a) each organelle- type's proximity to the cell body, to (3) characterize how these distributions change in a transgenic mouse model (5XFAD) of familial Alzheimer's disease (FAD) as a function of (b) the age of the mice & (c) gross brain region (hippocampus & isocortex) when compared to controls, & to (4) characterize how the colocalization of APP, Aßx, & ß-secretase changes within these organelles with respect to a, b, & c. Aim (1) will be achieved using coarse-grained molecular dynamics simulations of liposomes with encapsulated Aßx. Aims (2)-(4) will be achieved using fluorescence microscopy techniques. The proposed work is driven by the following Hypotheses: (1) a decreased pH will promote self-aggregation of Aßx & interactions of Aßx with the liposome walls, leading to disruption of the integrity of the liposome. (2) The pH distribution of neuronal organelles in all non-AD cases will exhibit a spatial gradient in which the organelles proximal to the cell body will be more acidic than distal organelles. (3) As the 5XFAD mice age, there will be (i) acidosis of the Golgi & early endosomes (EEs) & (ii) a statistically significant increase in th colocalization of Aßx & the ß-secretase enzyme in the Golgi & EEs as compared to the LEs & LYs in the AD model. Changes (i) & (ii) will both appear in neurons of the hippocampus before appearing in neurons of the isocortex. The proposed work seeks to bring the NIH's NIA one step closer to its goal of "understand[ing] the nature of aging & the aging process, & diseases & conditions associated with growing older" as it relates to AD, a ruthless, currently unstoppable, worldwide epidemic.