Table_3_An Integrated Pharmacology-Based Analysis for Antidepressant Mechanism of Chinese Herbal Formula Xiao-Yao-San.DOCX

Clinical studies and basic science experiments have widely demonstrated the antidepressant and anxiolytic effects of the herbal formula Xiao-Yao-San (XYS). However, the system mechanism of these effects has not been fully characterized. The present study conducted a comprehensive network pharmacological analysis of XYS and sorted all pharmacologically active components (149) through the TCMSP webserver. Then, all potential molecular targets (449) were predicted, of which there were 99 genes clearly related to depression. To further investigate the mechanism of antidepressant effects of XYS, a compound-depression targets (C-DTs) network was constructed, and Gene Ontology (GO) functional and KEGG pathway enrichment analyses were performed for the 99 targets. Enrichment results revealed that XYS could regulate multiple aspects of depression through these targets, related to metabolism, neuroendocrine function, and neuroimmunity. Prediction and analysis of protein–protein interactions resulted in selection of three hub genes (AKT1, TP53, and VEGFA). In addition, a total of seven ingredients from XYS could act on these hub genes and they were identified through ultra-high-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-Q/TOF-MS), including paeoniflorin, quercetin, luteolin, acacetin, aloe-emodin, Glyasperin C, kaempferol. Hereafter, we investigated the effects of paeoniflorin and its predicted target, the results suggest that it can reverse the neurotoxicity produced by CORT and could be a neuroprotective effect by promoting the phosphorylation of Akt. Overall, our research revealed the complicated antidepressant mechanism of XYS, and also provided a rational strategy for revealing the complex composition and function of Chinese herbal formula.

多项临床研究与基础科学实验已广泛证实，中药方剂逍遥散（Xiao-Yao-San, XYS）具备抗抑郁与抗焦虑活性。然而，其发挥此类功效的系统性作用机制尚未完全阐明。本研究针对逍遥散开展了全面的网络药理学分析，并通过TCMSP服务器筛选得到共计149个药理活性成分。随后，研究预测得到全部潜在分子靶点共449个，其中99个基因与抑郁症明确相关。为进一步阐明逍遥散的抗抑郁作用机制，本研究构建了化合物-抑郁症靶点（C-DTs）调控网络，并针对上述99个靶点开展了基因本体（Gene Ontology, GO）功能富集分析与KEGG通路富集分析。富集分析结果显示，逍遥散可通过上述靶点调控抑郁症发生发展的多个层面，涉及代谢、神经内分泌功能及神经免疫过程。通过蛋白质相互作用预测与分析，本研究筛选得到3个核心靶点基因（AKT1、TP53及VEGFA）。此外，本研究通过超高效液相色谱-四极杆飞行时间质谱（UPLC-Q/TOF-MS）鉴定得到逍遥散中共计7种可作用于上述核心靶点的活性成分，分别为芍药苷（paeoniflorin）、槲皮素（quercetin）、木犀草素（luteolin）、刺槐素（acacetin）、芦荟大黄素（aloe-emodin）、甘草素C（Glyasperin C）及山奈酚（kaempferol）。后续本研究针对芍药苷及其预测靶点开展了功能验证实验，结果显示芍药苷可逆转皮质酮（CORT）诱导的神经毒性，并通过促进Akt磷酸化发挥神经保护作用。综上，本研究阐明了逍遥散复杂的抗抑郁作用机制，同时为解析中药方剂的复杂组成与功能提供了一套科学可行的研究策略。