基于Caveolin-1通过NMDAR/ERK/CREB通路影响突触发育及重塑探讨固本健脑液对AD的作用及机制

Alzheimer's disease (AD) is a common multifactorial neurodegenerative disease, which has been confirmed that multiple anti-AD drugs for single target were invalid for AD. Traditional Chinese medicine has the advantage of multi-target pharmacological activity, therefore It is very important to seek the effective TCM treatment and classical prescription of AD. Caveolin-1, an attractive molecular target to augment synaptic plasticity in the hippocampus, may regulate multiple NMDAR-mediated signaling events involved in neuroprotection. In this sudy we aimed to explore the effect of Root-Securing and Brain-Fortifying Liquid (RSBFL) on the prevention and treatment of AD. Taking APP/PS1 animal model and N2a/APP cell model as the objects, the learning and memory ability and synaptic plasticity in AD mice dealed with RSBFL were observed, the expression Caveolin-1 and NMDAR-related molecules were tested by Western blot and RT-PCR, then the regulatory mechanism was verified through downregulating Caveolin-1 expression by cell transfection and with the use of NMDAR pathway inhibitor to in vitro, to clarify the mechanisms of RSBFL through activation of NMDAR/ERK/CREB signaling pathway via up-regulation of Caveolin-1, which could provide scientific basis for clinical medicine and lay the foundation for the development and research of classic prescription on AD.

阿尔茨海默病（AD）是常见的多因素性中枢神经退行性疾病，目前临床已证实针对单靶点的多种抗AD新药治疗无效，而中医药具有多靶点药理学活性的优势，因此寻求防治AD有效的中医治法和经典方药具有非常重要的意义。本项目拟围绕Caveolin-1有增强与神经发育和突触重塑密切相关的NMDAR信号通路功能的特点，深入探讨固本健脑液防治AD的作用。以双转基因AD小鼠及N2a/APP细胞为研究对象，首先观察固本健脑液对AD小鼠学习记忆能力及突触可塑性的影响，用Western blot和RT-PCR检测Caveolin-1和NMDAR通路相关分子表达，然后用细胞转染技术沉默Caveolin-1表达和用NMDAR通路抑制剂作用后在细胞水平验证其调控机制，阐明固本健脑液通过上调Caveolin-1表达活化NMDAR/ERK/CREB信号通路，进而促进神经发育及突触重塑达到神经保护作用，为中医防治AD提供理论依据。

阿尔茨海默病（AD）是常见的多因素性中枢神经退行性疾病，目前临床已证实针对单靶点的多种抗AD新药治疗无效，而中医药具有多靶点药理学活性的优势，因此寻求防治AD有效的中医治法和经典方药具有非常重要的意义。本项目在前期研究基础上基于Caveolin-1有增强与神经发育和突触重塑密切相关的NMDAR信号通路功能的特点，深入探讨固本健脑液防治AD的作用。本研究在系统阐述固本健脑法治则治法理论基础上，以双转基因AD小鼠及N2a/APP细胞为研究对象，首先观察固本健脑液对AD小鼠学习记忆能力及突触可塑性的影响，用Western blot和RT-PCR检测NMDAR通路相关分子表达，然后在细胞水平验证其调控机制，阐明固本健脑液通过调节NMDAR/ERK/CREB信号通路，进而促进神经发育及突触重塑达到神经保护作用，为中医防治AD提供理论依据。.在动物实验中，课题组以多奈哌齐为对照，运用固本健脑液对APP/PS1小鼠进行干预，水迷宫定位航行实验和空间探索实验结果表明固本健脑法能明显改善AD模型小鼠的学习记忆功能。WB、PCR、免疫组化、高尔基染色等检测发现，表明固本健脑液可提高 AD 模型小鼠皮质和海马NMDAR/ERK/CREB通路相关蛋白表达，增加突触的可塑性，并与减少Aβ1-42有关。.细胞实验部分，用固本健脑液提取物、多奈哌齐溶液、盐酸美金刚溶液分别干预各组细胞后，通过WB、PCR、ELISA等检测发现，固本健脑液提取物可降低APP695swe细胞培养基Aβ1-42含量，提高APP695swe细胞中NR2B、ERK1/2、CREB蛋白和基因的表达，逆转美金刚对APP695swe细胞NMDAR/ERK/CREB通路的抑制。可见，固本健脑液提取物可降低APP695swe细胞培养基Aβ1-42含量，提高APP695swe细胞中NR2B、ERK1/2、CREB蛋白和基因的表达，逆转美金刚对APP695swe细胞NMDAR/ERK/CREB通路的抑制。.以上研究证实，固本健脑液可有效防治AD，其机制可能与激活NMDAR/ERK/CREB通路，增加突触的可塑性，减少Aβ1-42有关。