网络药理学与实验证据：柴胡疏肝散的抗抑郁作用涉及 PI3K/AKT 信号通路。

OBJECTIVE: Chaihu Shugan San (CSS) is a common antidepressant prescription in traditional Chinese medicines. However, its active ingredients and mechanisms are unknown. The aim of this study was to explore the potential active ingredients and pharmacological mechanisms of CSS for the treatment of major depressive disorder (MDD). METHODS: Active compounds in CSS were screened using the Traditional Chinese Medicine Systems Pharmacology database. Compound-related targets were retrieved using the SwissTargetPrediction database. MDD-related targets were determined using DisGeNET, Therapeutic Target Database and DrugBank databases. The common targets of active compounds in CSS and MDD were retained to construct a compound-MDD target network. Then, functional enrichment analysis and protein-protein interaction analysis were performed to identify hub targets and explore the underlying molecular mechanisms. Finally, hub-targeted genes and pathways were validated by Western blotting and immunofluorescence using chronic unpredictable mild stress (CUMS) mice with or without CSS treatment. The affinities between the active compounds in CSS and hub-targeted genes were evaluated by molecular docking. RESULTS: Network pharmacology analysis revealed 24 potential targets for treatment of MDD by CSS. Functional enrichment analysis showed that PI3K/AKT signaling pathway was likely to be evidently affected by CSS in the treatment of MDD. In vivo experiments showed that CSS could improve depressive-like behaviors and promote neurogenesis in CUMS mice. Furthermore, CSS could increase phosphorylated (p) PI3K/PI3K and pAKT/AKT levels and decrease the pGSK3β/GSK3β level in the hippocampus of CUMS mice. The active compounds mainly included quercetin and luteolin, which showed good docking scores targeting the PI3K protein. CONCLUSION: This network pharmacological and experimental study highlights that the PI3K/AKT pathway is the potential mechanism by which CSS is involved in MDD treatment. Quercetin, luteolin, and kaempferol are probable active compounds in CSS, and these results might provide valuable guidance for further studies of MDD treatment.

目的：柴胡疏肝散（CSS）是一种常见的抗抑郁中药方剂。然而，其活性成分和作用机制尚不清楚。本研究旨在探讨 CSS 治疗重度抑郁症（MDD）的潜在活性成分和药理机制。

方法：使用中药系统药理学数据库筛选 CSS 的活性化合物。使用瑞士靶向预测数据库检索与化合物相关的靶点。使用 DisGeNET、治疗靶点数据库和 DrugBank 数据库确定 MDD 相关靶点。保留 CSS 活性化合物与 MDD 的共同靶点，构建化合物-MDD 靶点网络。然后进行功能富集分析和蛋白质-蛋白质相互作用分析，以鉴定关键靶点并探讨潜在的分子机制。最后，通过慢性不可预测性温和应激（CUMS）小鼠模型，结合 CSS 治疗，利用 Western blot 和免疫荧光技术验证关键靶向基因和通路。通过分子对接评估 CSS 中活性化合物与关键靶向基因的亲和力。

结果：网络药理学分析显示，CSS 治疗 MDD 的潜在靶点有 24 个。功能富集分析表明，PI3K/AKT 信号通路可能是 CSS 治疗 MDD 的重要靶点。体内实验表明，CSS 可改善 CUMS 小鼠的抑郁样行为，促进神经发生。此外，CSS 可增加 CUMS 小鼠海马组织中磷酸化（p）PI3K/PI3K 和 pAKT/AKT 水平，降低 pGSK3β/GSK3β 水平。活性化合物主要包括槲皮素和木犀草素，它们与 PI3K 蛋白的结合评分较好。

结论：本研究通过网络药理学和实验研究，强调了 PI3K/AKT 通路是 CSS 参与治疗 MDD 的潜在机制。槲皮素、木犀草素和山奈酚可能是 CSS 的活性化合物，这些结果可能为进一步研究 MDD 的治疗提供有价值的指导。