Astrocyte Activation, Dysfunction & Apoptosis in HAD

星形胶质细胞激活、功能障碍

• 批准号: 7016242
• 负责人: Dennis Larry Kolson
• 金额: $ 39.86万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-07-01 至 2010-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7016242
• 关键词: AIDS dementia complex; CD95 molecule; apoptosis; astrocytes; disease /disorder model; embryo /fetus; glutamates; human subject; laboratory rat; neurons; neuroprotectants; neurotransmitter transport; tissue /cell culture; tumor necrosis factor alpha

The cardinal pathological features of HIV-associated dementia (HAD) include astrogliosis/activation, macrophage/microglia (M/M) activation, synaptic damage, and apoptosis of neurons and astrocytes. The mechanisms of M/M activation in HAD remain uncertain, but factors released by HIV-activated M/M (glutamate, Fas-L, TNF-alpha, viral proteins, others) are likely responsible for astrocytic & neuronal injury through multiple mechanisms. Moreover, astrocyte activation and injury are critical in amplifying neuronal damage, most likely through impairment of astrocyte high-affinity glutamate scavenging and related neuroprotective functions. The mechanisms by which HIV/MM injure astrocvtes have received relatively little attention despite the evidence that this is critical in HAD pathogenesis. Our hypothesis is that HIV-activated M/M induce both astrocyte and neuronal injury and apoptosis as well as astrocyte activation and associated dysfunction; and that this exacerbates neuronal injury through loss of neuroprotective functions such as glutamate scavenging. We have developed in vitro models for neuronal and astrocytic apoptosis and dysfunction and have shown that mitochondrial-mediated (intrinsic) apoptosis triggered by HIV-M/M excitotoxins defines a major pathway by which HIV-M/M injure neurons. We also established a unique human astrocyte cell model with inducible trans-gene expression, and have modulated mitochondrial-mediated astrocyte apoptosis through Bcl-2 expression. Furthermore, we found that Fas-L, which is unregulated in HAD brain, suppresses glutamate transport in astrocytes. and that TNF-alpha suppresses glutamate transport in astrocytes and enhances transport in M/M. Finally, we have carried out in vivo studies using a novel single cell mRNA amplification/gene profiling approach to begin to define gene expression patterns of individual brain cells in HAD. Our goal is to utilize our in vitro models to define the pathways of astrocyte activation and associated dysfunction & apoptosis elicited by HIV/MM, and exploit our in vivo gene expression analysis approach to define the pathways activated in vivo in HAD. We will: 1) Identify the mechanisms and pathways of astrocyte activation and apoptosis induced by HIV-M/M and the role of Fas death receptors; 2) Determine the mechanisms of death receptor (Fas, TNF) modulation of high-affinity glutamate transport in astrocytes and M/M and the effects on neuronal survival; and 3) Define the in vivo pathways of astrocyte apoptosis and the patterns of activation regulating high-affinity glutamate transporter expression in HAD using our single-cell mRNA analysis to validate and extend in vitro findings. Understanding the mechanisms of astrocyte injury in HAD may identify additional targets for comprehensive neuroprotection in HAD.

HIV相关痴呆（HAD）的主要病理特征包括星形胶质细胞增生/激活、巨噬细胞/小胶质细胞（M/M）激活、突触损伤以及神经元和星形胶质细胞凋亡。 HAD 中 M/M 激活的机制仍不确定，但 HIV 激活的 M/M 释放的因子（谷氨酸、Fas-L、TNF-α、病毒蛋白等）可能通过多种机制导致星形胶质细胞和神经元损伤。此外，星形胶质细胞的激活和损伤对于放大神经元损伤至关重要，很可能 通过损害星形胶质细胞高亲和力谷氨酸清除和相关的神经保护功能。 HIV/MM 损伤星形细胞的机制相对较少受到关注，尽管有证据表明这在 HAD 发病机制中至关重要。我们的假设是，HIV 激活的 M/M 会诱导星形胶质细胞和神经元损伤和凋亡，以及星形胶质细胞激活和相关功能障碍；并且这会因谷氨酸清除等神经保护功能的丧失而加剧神经元损伤。我们开发了神经元和星形细胞凋亡和功能障碍的体外模型，并表明由 HIV-M/M 兴奋性毒素触发的线粒体介导的（内在的）细胞凋亡定义了一个主要途径，通过该途径 HIV-M/M 损伤神经元。我们还建立了具有可诱导转基因表达的独特的人类星形胶质细胞模型，并通过 Bcl-2 表达调节线粒体介导的星形胶质细胞凋亡。此外，我们发现在 HAD 大脑中不受调节的 Fas-L 会抑制星形胶质细胞中的谷氨酸转运。 TNF-α 抑制星形胶质细胞中的谷氨酸转运并增强 M/M 中的转运。最后我们在vivo进行了 研究使用新型单细胞 mRNA 扩增/基因分析方法开始定义 HAD 中单个脑细胞的基因表达模式。我们的目标是利用我们的体外模型来定义星形胶质细胞激活以及由 HIV/MM 引起的相关功能障碍和细胞凋亡的途径，并利用我们的体内基因表达分析方法来定义 HAD 体内激活的途径。我们将： 1）明确HIV-M/M诱导星形胶质细胞活化和凋亡的机制和途径以及Fas死亡受体的作用； 2）确定死亡受体（Fas、TNF）调节星形胶质细胞和M/M中高亲和力谷氨酸转运的机制以及对神经元存活的影响； 3) 使用我们的单细胞 mRNA 分析来验证和扩展体外研究结果，定义星形胶质细胞凋亡的体内途径以及调节 HAD 中高亲和力谷氨酸转运蛋白表达的激活模式。了解 HAD 中星形胶质细胞损伤的机制可能会确定其他目标 HAD 的全面神经保护。