JNK/ERK1/2信号转导通路对受体介导神经细胞内化β淀粉样蛋白及其作用的调控机制

It has been known that intracellular deposition of beta amyloid protein (Aβ) is earlier than extracellular one, and an important factor of abnormal neural activity and apoptosis in Alzheimer’s disease (AD). MAPK signal transduction pathways is activated in AD. Our previous study found that p38MAPK has a role in the regulation of internalization of Aβ. The MAPK signal transduction pathway including three subfamilies p38 MAPK, JNK and ERK1/2 all are involved in the internalization of protein related events. However, whether JNK and ERK1/2 signal transduction pathways are also involved in the regulation of the receptor-mediated internalization of Aβ and its effects is little-known already. This study will be on the basis of previous work, focus on the regulation of JNK/ERK1/2 signal transduction pathway on the receptor-mediated internalization of Aβ and its effects in neuron. First of all, using morphological method, RT-PCR, ELISA combined with behavioral, electrophysiological (MED planar microelectrode array record LTP/LTD) technology to observe and analyze the Aβ internalized into neural cell types, the quantity of Aβ deposition, the relationship between the expression of low-density lipoprotein receptor-related protein-1(LRP1), alpha 7 nicotinic acetylcholine receptor (α7nAChR ) and JNK/ ERK1/2 signal transduction pathway. Meanwhile, to analysis apoptotic related events, the changes of synaptic plasticity and learning and memory in mice. Secondly, the immunoprecipitation, siRNA and inhibitor of JNK/ERK1/2 are emploied to investigate the effects of JNK/ERK1/2 signal transduction pathway on the regulation of the internalization of Aβ and its effects and verify the related receptor mediated internalization of Aβ. To elucidate the regulation of the internalization of Aβ and its effects by JNK/ERK1/2 signal transduction pathway and reveal the pathogenesis of AD, and provide new ideas and targets for the treatment of AD.

已知细胞内Aβ沉积先于细胞外，是导致AD神经活动异常和细胞凋亡的重要因素。我们前期的研究发现p38MAPK对细胞内化Aβ具有调控作用，在MAPK家族中p38、JNK、ERK1/2都参与了和蛋白质内化相关事件，而后两者是否也参与细胞内化Aβ鲜为人知。本课题探索JNK/ERK1/2对受体介导神经细胞内化Aβ及其作用调控机制。首先采用形态学、RT-PCR、ELISA结合行为学、电生理学等技术观察分析Aβ内化入神经细胞类型、Aβ沉积量，Aβ内化相关受体LRP1、α7nAChR与ERK1/2/JNK信号通路间的关系，同时分析细胞凋亡相关事件、神经突触可塑性、学习记忆力；其次应用免疫共沉淀、RNAi技术结合通路阻断剂分析ERK1/2/JNK对LRP1、α7nACh介导神经细胞内化Aβ及其作用影响，阐明JNK/ERK1/2对神经细胞内化Aβ及其作用调控机制，为揭示AD发病机制及AD治疗提供新思路和靶点。

阿尔茨海默病（Alzheimer’s disease, AD）是一种渐进性神经退行性疾病，临床症状以近期记忆力减退、认知功能障碍、行为异常和社交障碍为主。最新报告AD为威胁老年人健康的“第五大杀手”。由于AD发病的复杂性，其发病机制尚未阐清。. 本课题综合运用神经科学研究技术和方法，进一步探索MAPKs通路对受体介导神经细胞内化β淀粉样蛋白及其作用的调控机制。主要研究内容涉及到五个方面：内化Aβ的神经细胞种类、在亚细胞结构的定位，LRP1、α7nAChR表达及其与Aβ共定位及相互作用；神经细胞内化Aβ时细胞凋亡相关事件与MAPKs及LRP1/α7nAChR相关性及其神经活动功能变化；细胞内化Aβ与LRP1、α7nACh受体、MAPKs信号通路间的关系；MAPKs 信号转导通路对LRP1、α7nAChR介导Aβ内化及其作用的调控。.结果显示：Aβ能够被GABA能神经元和ChAT阳性神经元内化，也可被星形胶质细胞内化；Aβ被内化后主要定位在线粒体、溶酶体，内质网中有少量沉积；Aβ1-42寡聚体（oAβ1-42）神经毒性作用可产生氧化应激和炎症，引起细胞凋亡，导致学习记忆障碍。JNK信号通路未参与细胞内化Aβ1-42，而p38和ERK1/2通过影响α7nAChR和LRP1的表达而调控细胞内化Aβ1-42。激活α7nAChR，阻断p38或JNK改善Aβ引起的氧化应激，并减轻炎症，进而减少Aβ引起的细胞凋亡，上调学习记忆相关蛋白的表达，最终改善Aβ引起的空间记忆障碍，阻断ERK信号通路对Aβ引起的神经毒性作用无改善作用。小鼠海马脑区α7nAChR与LRP1蛋白表达水平与小鼠的空间记忆能力显著相关。.这一研究成果为阐明AD发病机制提供了重要理论和实验数据，为临床有效治疗AD提供了新靶点和思路。