星形胶质细胞“Aβ/PirB-mTOR-EAATs”通路抑制Glu摄取促进AD发展的作用研究

The excitotoxic neural death induced by the accumulation of excessive glutamate in synaptic cleft is one of the major pathogenesis of Alzheimer’s Disease. The residual extracellular glutamate is substantially removed by the glutamate transporters (EAATs) on astrocytes, while the regulatory mechanism of EAATs is barely clarified. Preliminary experiments and previous work of our team has indicated that PirB（paired immunoglobulin-like receptor B, a highly affinitive receptor of Aβ）, which participates in synaptic plasticity, could down-regulate EAATs expression by inhibiting mTOR activation. In this study the primary cultured neuronal cells is going to be transfected by lentiviral vectors to up-regulate or silencing PirB expression, or treated with soluble PirB extracellular fragments before Aβ stimulation, and the expression of EAATs, the uptake of glutamate and neural death will be determined by molecular biological, pharmacological and morphological methods; the mice which are conditionally knocked out PirB on astrocytes and the APP/PS1 mice of AD mice model will be hybridized to generate the astrocytic PirB knocking out APP/PS1 mice, the changes of the expression of EAATs, the activation of mTOR, the neural death as well as the cognitive behaviors of the mice were detected. This study aims to clarify the mechanism of the role of astrocytic “Aβ/PirB-mTOR-EAATs” signaling mediated excitotoxicity in AD pathogenesis. The great outstanding of this mechanism will provide some evidence for the treatment and prevention of AD.

突触间隙谷氨酸摄取障碍引起的神经元兴奋性毒性死亡是AD的重要发病机制。谷氨酸主要由星形胶质细胞（As）的谷氨酸转运体（EAATs）摄取，但EAATs表达的调节机制鲜有研究。我们预实验和前期工作表明在As细胞中，参与突触可塑性调节的PirB分子（一种Aβ高亲和力受体）可能通过抑制mTOR下调EAATs表达。本研究拟培养原代神经细胞，用慢病毒转染上调或沉默PirB，或采用可溶性PirB胞外段抑制内源性PirB，其后给予Aβ处理，用分子生物学、药理学、形态学等方法检测EAATs表达、谷氨酸摄取及神经元凋亡的改变；并将敲除As细胞PirB小鼠与APP/PS1转基因AD小鼠杂交，观察敲除As细胞PirB的APP/PS1小鼠EAATs表达、mTOR激活、神经元凋亡和认知行为改变，阐明As细胞“Aβ/PirB-mTOR-EAATs”介导神经元兴奋性毒性参与AD发展的机制，为AD的治疗和预防提供依据。

突触间隙谷氨酸摄取障碍引起的神经元兴奋性毒性死亡是AD的重要发病机制。谷氨酸主要由星形胶质细胞（As）的谷氨酸转运体（EAATs）摄取，但EAATs表达的调节机制鲜有研究。我们预实验和前期工作表明在As细胞中，参与突触可塑性调节的PirB分子（一种Aβ高亲和力受体）可能通过抑制mTOR下调EAATs表达。本研究培养了原代神经细胞，用慢病毒转染上调或沉默PirB，或采用可溶性PirB胞外段抑制内源性PirB，其后给予Aβ处理，用分子生物学、药理学、形态学等方法检测EAATs表达、谷氨酸摄取及神经元凋亡的改变；并将敲除As细胞PirB小鼠海马立体定位注射Aβ寡聚体，观察敲除As细胞PirB的小鼠EAATs表达、mTOR激活、 神经元凋亡和认知行为改变，阐明As细胞“Aβ/PirB-mTOR-EAATs”介导神经元兴奋性毒性参与AD发展的机制，为AD的治疗和预防提供依据。