Effects of opiates on neurons and their impact on HIV neuropathology

项目概要/摘要 HIV相关的神经认知障碍（HAND）在病毒抑制患者中持续存在。手是一个 异质性疾病，其特征在于慢性低水平炎症、兴奋性毒性、 神经毒性HIV蛋白，APP加工改变，以及其他加剧神经元结构和 功能阿片类药物的使用在HIV+患者中很常见，并被认为是导致HAND的原因， 仍然存在争议。上一个资助期的研究表明，吗啡可以通过以下方式增强HAND： 通过一种新型的铁依赖性途径改变APP加工。已知有毒APP裂解产物 减少大脑几个区域的树突棘，这是学习和记忆的关键介质。 令人惊讶的是，APP裂解产物对前额叶皮层（PFC）树突棘的影响， 这是一个对人力资源和发展至关重要的领域，但尚未得到深入研究。该项目将阐明 改变吗啡和艾滋病毒诱导的PCF神经元缺陷的APP加工。研究将揭开 HIV和Aβ蛋白相互作用，并确定吗啡是否有助于APP的改变 处理和刺在手中。目的1的研究将集中在Aβ寡聚体对树突状细胞的影响， PFC神经元中的棘结构和功能，在存在和不存在HIV神经毒素或吗啡的情况下。 这些体外和体内研究将确定树突棘密度/形态/转换的有意义的变化 在PFC神经元，并提供了一个全面的评估外源性增加Aβ的行动，在这个关键的大脑区域（ 内源性Aβ的作用将在目的3中进一步测试）。目标2的主要目标是确定 其中μ-阿片类药物可以通过调节神经元铁来影响淀粉样蛋白的生成。这些机械研究 是基于我们最近的发现，表明铁在吗啡介导的树突棘减少中的作用 大脑皮层神经元他们还得到了以下发现的支持，即内体脱酸导致细胞内蛋白质的增加。 胞浆Fe ~（2+）和Aβ。重要的是，胞质Fe 2+调节APP表达，而内溶酶体pH 调节蛋白质降解。在aim 3中，我们将采用新产生的分子工具， 加工到α-裂解途径或阻止APP被β-分泌酶裂解。经过功能测试后， 在原代培养物中，最有希望的构建体将在HAND动物模型的PFC神经元中表达， 确定它们在HIV蛋白/吗啡存在下如何有效地降低Aβ水平，以及Aβ降低是否 有助于PFC认知任务的恢复。这一建议将阐明铁依赖性机制， 艾滋病毒+/阿片类药物滥用受试者的认知能力加速下降，这与ART- 接受治疗的患者寿命更长。

Project Summary/Abstract HIV-associated neurocognitive disorders (HAND) persist in virally suppressed patients. HAND is a heterogeneous disease that is characterized by chronic low-level inflammation, excitotoxicity, presence of neurotoxic HIV proteins, altered APP processing, and other factors that aggravate neuronal structure and function. Opioid use is common among HIV+ patients and thought to contribute to HAND, although this remains controversial. Studies from the previous funding period suggest that morphine can augment HAND by altering APP processing through a novel, iron-dependent pathway. Toxic APP cleavage products are known to reduce dendritic spines in several brain areas, which are critical mediators of learning and memory. Surprisingly, the effects of APP cleavage products on dendritic spines of the prefrontal cortex (PFC), which is an area of critical importance to HAND, have not been studied in depth. This project will elucidate the role of altered APP processing in morphine and HIV-induced neuronal deficits in the PCF. Studies will unravel the interaction between HIV and Aβ proteins and determine whether morphine can contribute to alteration of APP processing and spines in HAND. Research in aim 1 will concentrate on the effect of Aβ oligomers on dendritic spine structure and function in PFC neurons, in the presence and absence of HIV neurotoxins or morphine. These in vitro and in vivo studies will define meaningful changes in dendritic spine density/morphology/turnover in PFC neurons, and provide a full assessment of exogenously added Aβ actions in this critical brain area (the role of endogenous Aβ will be further tested in aim 3). The main goal of aim 2 is to establish the extent to which µ-opioids can affect amyloidogenesis through modulation of neuronal iron. These mechanistic studies are based on our recent discoveries suggesting a role for iron in morphine-mediated dendritic spine reduction in cortical neurons. They are also supported by the finding that endosome deacidification leads to increase of cytosolic Fe2+ and Aβ. Importantly, cytosolic Fe2+ regulates APP expression whilst endolysosomal pH modulates protein degradation. In aim 3, we will employ newly generated molecular tools able to shift APP processing to the α-cleavage pathway or prevent APP cleavage by β-secretases. After functional testing in primary cultures, the most promising constructs will be expressed in PFC neurons of HAND animal models to determine how effectively they reduce Aβ levels in the presence of HIV proteins/morphine and if Aβ reduction contributes to recovery of PFC cognitive tasks. This proposal will shed light on iron-dependent mechanisms of accelerated cognitive decline in HIV+/opiate abusing subjects, which is becoming increasingly relevant as ART- treated patients live longer.