aFGF改构体调控GSK3β磷酸化及其靶标分子治疗AD的分子机制

Alzheimer's disease is a severely destructive neuro degenerative disease characterized by hippocampus and cortex neurons degeneration .Studies have shown that GSK3β is directly related to the aggregation of Aβ and tau protein abnormal phosphorylation, which plays an important role in the development of alzheimer's disease (AD). Based on the powerful potency in repairing the nerve injury and protecting from neuron apoptosis acidic fibroblast growth factor (aFGF), we firstly reported that aFGF delivered by cell-penetrating peptide TAT can effectively penetrate the blood brain barrier, improve the cognitive, learning and memory ability of AD mice, and decrease the formation of senile plaque (SP) and neurofibrillary tangles (NFT) in hippocampus and cortical; Preliminary mechanism research shows that aFGF is involved in the regulation of GSK3β phosphorylation, and then GSK3β regulates Aβ generation, tau protein phosphorylation and neuron apoptosis. In this project, our studies will entry from the regulation of aFGF on the GSK3β phosphorylation. Co-immunoprecipitation assay and iTRAQ labeling quantitative proteomic technology are used to identify the key target proteins interacting with GSK3β. Then, gene overexpression and known-down technology are employed to analyze the mechanisms of these interactions, and to systematically identify the regulation of aFGF medicated by GSK3β on the Aβ metabolism, Tau phosphorylation, and the signal pathway of neuronal apoptosis. This study will provide the experimental evidence and rationale to the pathogenesis of AD and other neurodegenerative diseases, and the development of new therapeutic targets for AD treatment drugs.

阿尔茨海默病（AD）以神经退行性病变为病理特征，发病机制尚未明确。糖元合成酶3β（GSK3β）参与Aβ生成和Tau磷酸化，对AD发生发展有重要的调节作用。基于酸性成纤维细胞生长因子（aFGF）的损伤修复、神经营养和抗神经元凋亡等功能，我们首次报道了穿膜肽能有效介导aFGF穿过血脑屏障，改善AD模型鼠认知和学习记忆能力，减少海马与皮质中老年斑（SP）和神经原纤维缠绕（NFT）的形成；初步机制研究显示aFGF参与GSK3β磷酸化的调节，进而调节Aβ生成、Tau磷酸化及神经元凋亡。本项目以aFGF调控GSK3β磷酸化为切入点，采用免疫共沉淀和iTRAQ蛋白组学技术，筛选出与GSK3β特异性结合的关键靶蛋白，应用基因过表达和干扰等技术研究相互作用机制，系统地阐明aFGF通过GSK3β调控Aβ代谢、Tau磷酸化和神经元凋亡的信号通路，为AD发病机制研究和新靶点治疗药物开发提供理论基础和实验依据。

阿尔茨海默病（AD）以神经退行性病变为病理特征，发病机制尚未明确。酸性成纤维细胞生长因子（aFGF）具有损伤修复、神经营养和抗神经元凋亡等功能，但由于FGF类大分子药物分子量大无法穿过血脑屏障直接入脑，制约了其在神经退行性疾病方面的应用。.本研究采用鼻腔给药、脂质体包裹和Tat-PTD蛋白融合技术进行联用研究并建立了新型的给药系统，解决了生物大分子药物穿过血脑屏障入脑的瓶颈问题。课题组采用滴鼻给药脂质体Tat-haFGF能显著提高haFGF药物进入脑内的浓度，对APP/PS1模型动物的神经退行性病变具有显著的治疗效果。同时体外用Aβ损伤大鼠原代皮质神经元作为AD细胞模型，发现Tat-haFGF14-154 同样可以拮抗Aβ引起的神经元损伤。通过体内外实验对Tat-haFGF14-154的神经保护机制进行研究发现，ADMA10 是Tat-haFGF14-154 在AD模型中发挥神经保护作用的重要靶标分子，Tat-haFGF14-154 通过PI3K-CREB-IRE1α/XBP1信号通路促进ADAM10的转录表达，从而发挥神经保护作用。同时，用过表达APP的细胞株N2a-APP作为细胞模型，通过免疫共沉淀、蛋白组学等技术，发现Tat-haFGF14-154可通过CRMP蛋白调控GSK3β的磷酸化进而起到神经保护作用。.本研究为针对多靶点治疗AD的新型药物开发和揭示AD等神经退行性疾病的发病机制提供重要的理论基础和实验依据，为神经营养类药物在神经退行性疾病治疗方面的用药方法提供参考。