Table_7_The Food Additive β-Caryophyllene Exerts Its Neuroprotective Effects Through the JAK2-STAT3-BACE1 Pathway.xlsx

Despite extensive research on Alzheimer’s disease (AD), its diagnosis and treatment remain challenging, and no effective therapies are currently available. Amyloid β (Aβ) extracellular plaques and intracellular neurofibrillary tangles are the histological characteristics of AD that have been directly linked to neuropathological events such as synaptic and neuronal cell loss. In this study, we explored whether the “JAK2-STAT3-BACE1” pathway is involved in neuroprotection conferred by the food flavouring agent β-caryophyllene (BCP). PC-12 cells with overexpressed amyloid-β protein precursor (APP) were utilised to construct an AD model in vitro, which was then split into four groups, namely control, empty vector, APP overexpression, and BCP (5, 10, and 20 μM). CCK-8 was used to evaluate cell viability, immunofluorescence was utilised to examine synaptic morphology, and quantitative real-time polymerase chain reaction and western blot were used to examine gene and protein expression levels. The relative expression levels of JAK2, STAT3, and BACE1 mRNA in the transfected PC-12 cells were found to be significantly upregulated. The cell morphology altered dramatically 72 h after transfection, becoming rounder, with a decrease in cell number. BCP exhibited the potential to dramatically increase PC-12 cell viability while protecting cell morphology. BCP inhibited APP, JAK2, STAT3, BACE1 mRNA and BACE1 protein overexpression, as well as JAK2 and STAT3 hyperphosphorylation. Molecular docking simulated the docking of BCP with JAK2, STAT3, BACE1, CB2. And JAK2 was found to be the most stable protein. In conclusion, inhibition of the “JAK2-STAT3-BACE1” signalling pathway may be one of the mechanisms through which BCP protects neurons and antagonises Aβ’s neurotoxicity.

尽管针对阿尔茨海默病（Alzheimer’s disease, AD）已开展了大量研究，但其诊断与治疗仍面临诸多挑战，目前尚无有效的治疗手段可供使用。淀粉样β蛋白（Amyloid β, Aβ）细胞外斑块与细胞内神经原纤维缠结是阿尔茨海默病的典型组织病理学特征，二者与突触丢失、神经元细胞死亡等神经病理事件直接相关。本研究旨在探究“JAK2-STAT3-BACE1”通路是否参与食品风味剂β-石竹烯（β-caryophyllene, BCP）所介导的神经保护作用。本研究使用过表达淀粉样前体蛋白（amyloid-β protein precursor, APP）的PC-12细胞构建体外AD模型，随后将模型分为四组：对照组、空载体组、APP过表达组以及不同浓度BCP处理组（5、10和20 μM）。采用细胞计数试剂盒-8（CCK-8）评估细胞活力，利用免疫荧光技术检测突触形态，通过实时定量聚合酶链反应（quantitative real-time polymerase chain reaction, qRT-PCR）与蛋白质印迹（western blot）检测基因与蛋白的表达水平。研究结果显示，转染后的PC-12细胞中JAK2、STAT3及BACE1的mRNA相对表达水平显著上调；转染72小时后，细胞形态发生显著改变，形态趋于圆润，细胞数量有所减少。BCP可显著提升PC-12细胞的活力，同时对细胞形态起到保护作用。BCP能够抑制APP、JAK2、STAT3、BACE1的mRNA以及BACE1蛋白的过表达，同时抑制JAK2与STAT3的过度磷酸化。分子对接模拟了BCP与JAK2、STAT3、BACE1、CB2的对接作用，且发现JAK2是对接稳定性最高的蛋白。综上，抑制“JAK2-STAT3-BACE1”信号通路可能是BCP发挥神经元保护作用、拮抗Aβ神经毒性的潜在机制之一。