Liu Shuo, Xu Luting, Shen Yan, Wang Liuying, Lai Xiaoxiao, Hu Haiyan
The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, 109 Xue Yuan Xi Road, Lu Cheng District, Wenzhou, 325000, China; The Second Clinical College, Wenzhou Medical University, Wenzhou, 325003, China.
The Second Affiliated Hospital and Yuying Children's Hospital of Wenzhou Medical University, 109 Xue Yuan Xi Road, Lu Cheng District, Wenzhou, 325000, China; The Second Clinical College, Wenzhou Medical University, Wenzhou, 325003, China.
J Ethnopharmacol. 2024 Jan 10;318(Pt B):117031. doi: 10.1016/j.jep.2023.117031. Epub 2023 Aug 12.
Alzheimer's disease (AD) belongs to the category of "senile dementia" in traditional Chinese medicine. AD is associated with brain emptiness or collaterals blocked by phlegm-heat. "Fumanjian" from Jingyue Quanshu treats dementia by promoting qi circulation, alleviating depression, eliminating turbidity, cultivating positivity, and dispelling evil spirits. Qingxin Kaiqiao Fang (QKF), derived from Fumanjian, is effective in treating AD owing to previously mentioned clinical effects. Elucidating the mechanism(s) of action of QKF on AD associated with phlegm-heat may be beneficial for therapeutic management; however, further research is needed.
This study aimed to determine the role of the PI3K/Akt pathway in AD, especially the specific effector protein involved, and explore the efficacy of QKF in treating AD by modulating the PI3K/Akt signal.
High-performance liquid chromatography-Q-orbitrap-mass spectrometry was used to analyze the chemical components of QKF. Subsequently, APP/PS1 double-transgenic mice were used for behavioral tests, and hematoxylin-eosin and Nissl staining were used to assess the neuroprotective and cognitive effects of QKF. Cerebrospinal fluid pharmacology was used in in vitro validation, and Aβ was used to induce PC12 cells to establish the AD cell model. Various methods, including immunohistochemistry, Western blotting, quantitative real-time polymerase chain reaction, morphological assay, cell counting kit-8(CCK-8) assay, and terminal deoxynucleotide transferase (TdT)-mediated dUTP nick-end labeling (TUNEL)staining, were used to evaluate the effect of QKF on Tau hyperphosphorylation and anti-apoptosis. These methods also assessed the influence of QKF on the PI3K/Akt/GSK3β pathway involving the mRNA and protein expressions. Finally, the inhibitor - LY294002 was used for reverse validation.
We identified 295 chemical components in the water extract of QKF.QKF improved spatial cognition and learning memory in APP/PS1 mice, protected PC12 cell morphology, improved cell survival, reduced Aβ-induced apoptosis, and inhibited the hyperphosphorylation of Tau protein via the PI3k/Akt/GSK3β signaling pathway. Furthermore, this protective effect of QKF was reduced by LY294002 in vitro.
QKF can improve spatial cognition, learning, and memory abilities in APP/PS1 mice and protect PC12 cells. Decreasing the Tau hyperphosphorylation in AD exhibits curative efficacy on AD via the PI3K/Akt/GSK3β pathway in vitro and in vivo.
阿尔茨海默病（AD）在中医中属于“老年痴呆”范畴。AD与脑髓空虚或痰热阻滞经络有关。《景岳全书》中的“扶满健”通过理气解郁、化痰开窍、培本扶正、驱邪外出治疗痴呆。源自“扶满健”的清心开窍方（QKF），因其上述临床疗效，对治疗AD有效。阐明QKF对痰热型AD的作用机制可能有助于治疗管理；然而，仍需进一步研究。
本研究旨在确定PI3K/Akt信号通路在AD中的作用，特别是涉及的特定效应蛋白，并探讨QKF通过调节PI3K/Akt信号治疗AD的疗效。
采用高效液相色谱-Q-轨道阱-质谱联用技术分析QKF的化学成分。随后，将APP/PS1双转基因小鼠用于行为测试，并用苏木精-伊红染色和尼氏染色评估QKF的神经保护和认知作用。采用脑脊液药理学进行体外验证，并用Aβ诱导PC12细胞建立AD细胞模型。采用免疫组织化学、蛋白质免疫印迹法、实时定量聚合酶链反应、形态学分析、细胞计数试剂盒-8（CCK-8）检测法和末端脱氧核苷酸转移酶（TdT）介导的dUTP缺口末端标记（TUNEL）染色等多种方法，评估QKF对Tau蛋白过度磷酸化和抗凋亡的影响。这些方法还评估了QKF对涉及mRNA和蛋白表达的PI3K/Akt/GSK3β信号通路的影响。最后，使用抑制剂LY294002进行反向验证。
我们在QKF水提取物中鉴定出295种化学成分。QKF改善了APP/PS1小鼠的空间认知和学习记忆能力，保护了PC12细胞形态，提高了细胞存活率，减少了Aβ诱导的细胞凋亡，并通过PI3k/Akt/GSK3β信号通路抑制了Tau蛋白的过度磷酸化。此外，LY294002在体外降低了QKF的这种保护作用。
QKF可改善APP/PS1小鼠的空间认知、学习和记忆能力，并保护PC12细胞。降低AD中Tau蛋白的过度磷酸化在体外和体内通过PI3K/Akt/GSK3β信号通路对AD具有治疗作用。
