HIV-1Tat抑制EAAT-2表达参与HAD发生的分子机制

With the introduction of highly active anti-retroviral therapy has been successful to reduce progression of acquired immunodeficiency syndrome (AIDS), the lifespan of human immunodeficiency virus (HIV) carriers has been dramatically elongated. However, this also leads to an increased incidence of HIV-associated dementia (HAD). Our previous study prompts that decreased excitatory amino acid transporter 2 (EAAT-2) on the astrocytes accompanied with high levels of glutamate in the synaptic cleft is a key factor responsible for neuronal damage in the process of HAD. However, the underlying molecular mechanism remains unclear. In our preliminary study, we found that HIV-1Tat can upregulate the expression of the astrocyte elevated gene-1 (AEG-1) and downregulate EAAT-2 expression. In addition, we found that p38MAPK was highly phosphorylate upon overexpression of AEG-1 in mouse astrocytes. Furthermore, HIV-1Tat treatments resulted in excessive interleukin-lβ (IL-1β) generation in microglia. Thus, we speculate that HIV-1Tat downregulates EAAT-2 expression through at least two mechanisms in HAD: on one hand, HIV-1Tat may increase astrocyte AEG-1 expression and thus downregulate the expression of EAAT-2 through the p38-MAPK signaling pathway. On the other hand, HIV-1Tat may activate microglia to release excessive IL-1β, resulting in the downregulation of EAAT-2. Here, We will employ laser microdissection, ChIP, and other techniques to elucidate the molecular signaling networks involved in HIV-1Tat inhibitting EAAT-2 expression through AEG-1 and/or IL-1β, using SHIV infected macaque, clinical autopsy specimen and primary cultured astrocyte models. These studies will not only strengthen our understanding of the pathogenesis of HAD, but also identify new molecular targets for HAD treatment.

星形胶质细胞EAAT-2表达低下是导致HIV相关性痴呆（HAD）中胞外谷氨酸浓度过高诱导神经元凋亡的关键因素，然而其机制尚未阐明。我们发现HIV-1Tat可上调AEG-1，下调EAAT-2表达，并且AEG-1过表达的小鼠星形胶质细胞株中p38MAPK高度磷酸化；HIV-1Tat可使小胶质细胞产生过量IL-1β进而下调EAAT-2表达。由此我们推测HIV-1Tat至少通过以下两种机制下调EAAT-2表达，参与HAD的发生：①HIV-1Tat上调AEG-1表达，进而通过p38-MAPK等信号通路下调EAAT-2表达。②HIV-1Tat通过活化小胶质细胞释放过量的IL-1β间接调节EAAT-2表达。本项目拟从SHIV感染恒河猴动物实验、临床尸检标本分析、细胞模型三个层面，使用激光纤维切割、ChIP等技术，阐明HIV-1Tat抑制EAAT-2表达的分子机制，确定其调控的信号分子网络。

星形胶质细胞谷氨酸转运蛋白2（excitatory amino acid transporter-2，EAAT-2）表达低下是导致人类免疫缺陷病毒相关性痴呆 (human immunodeficiency virus-associated dementia，HAD)中胞外谷氨酸浓度过高诱导神经元凋亡的关键因素，然而其机制尚未阐明。我们使用11只SHIV感染的恒河猴动物模型探讨HIV-1Tat对EAAT-2的作用及其分子机制。我们发现，在SHIV感染的恒河猴大脑皮层中水通道蛋4（AQP4）表达减少，且AQP4表达减少与神经元和胶质细胞凋亡、及其HAD的发生有关。.我们进一步研究发现，在SHIV感染的恒河猴大脑皮层中星形胶质细胞高表达基因-1（Astrocyte-Elevated Gene-1, AEG-1）的表达增加，统计分析显示AEG-1阳性细胞数量与EAAT-2阳性面积呈负相关，提示AEG-1过表达与EAAT-2表达低下相关。.我们进一步使用细胞模型探讨EAAT-2表达低下的分子机制。我们使用HIV-1Tat 处理U87细胞后，Waster Blot 显示EAAT-2表达减少，提示HIV-1Tat可以下调EAAT-2的表达。进一步的研究结果证实HIV-1Tat可以通过PI3-K信号通路上调AEG-1的表达，而上调的AEG-1则通过NF-kB信号通路上调阴阳-1（Yin Yang-1,YY-1）进而起到抑制EAAT-2的作用。EAAT-2表达减少，突触间隙谷氨酸浓度过高或持续激活谷氨酸受体导致Ca2＋内流而引起神经元损伤。我们的结果将为HAD提供新的治疗靶点。

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