开心散抗抑郁作用的靶点群发现及分子机制研究  

Anti-depression targets and mechanism study of Kaixin San

作　　者：杨倬 卓芳芳 张贵民 孙成宏 屠鹏飞[1] 姚景春 曾克武[1] YANG Zhuo;ZHUO Fang-fang;ZHANG Gui-min;SUN Cheng-hong;TU Peng-fei;YAO Jing-chun;ZENG Ke-wu(State Key Laboratory of Natural and Biomimetic Drugs,School of Pharmaceutical Sciences,Peking University,Beijing 100191,China;State Key Laboratory of Generic Manufacture Technology of Chinese Traditional Medicine,Lunan Pharmaceutical Group Co.,Ltd.,Linyi 276006,China)

机构地区：[1]北京大学药学院天然药物及仿生药物国家重点实验室,北京100191 [2]鲁南制药集团股份有限公司中药制药共性技术国家重点实验室,山东临沂276006

出　　处：《中国中药杂志》2023年第2期472-480,共9页China Journal of Chinese Materia Medica

基　　金：国家自然科学基金项目(81973505,82174008);中央高校基本科研业务费专项。

摘　　要：基于靶点“钩钓”策略鉴定开心散在脑组织中的抗抑郁靶点群,探索靶点群相关药理学信号通路,诠释开心散抗抑郁的分子机制。通过慢性不可预见性温和应激(CUMS)方式建立Balb/c小鼠抑郁模型,以糖水偏好及强迫游泳行为学实验评价开心散抗抑郁作用;进而利用光交联反应将开心散药效组分键合到磁性微球表面,形成开心散成分芯片,从抑郁小鼠脑皮层、丘脑、海马组织中捕获直接作用靶点蛋白,并通过液相色谱高分辨质谱联用(LC-MS/MS)进行鉴定;利用Cytoscape软件进行靶点蛋白通路富集分析,同时通过免疫组化和蛋白免疫印记等方法对可能的靶点生物学功能进行验证。研究发现,开心散对抑郁小鼠的行为学指标有明显改善作用;靶点鉴定实验发现开心散药效组分在皮层、丘脑和海马组织中的潜在作用靶点分别有64、91和44个,靶点功能主要与加压素调控的水重吸收、沙门菌感染、甲状腺激素合成等信号通路相关。免疫组化和免疫印记等分析表明,开心散上调了抑郁小鼠脑皮层中加压素(AVP)的表达,上调了小鼠丘脑中热休克蛋白60(HSP60)、细胞色素C氧化酶4(COX4)、促甲状腺激素释放激素(TRH)的表达,并下调了丘脑中的肿瘤坏死因子α(TNF-α)及NF-κB p65的表达。因此,开心散可能调控了皮层中的加压素信号通路,以及丘脑中的能量代谢、甲状腺激素、炎症反应等信号通路发挥抗抑郁作用,而对海马影响较弱。This study identified the anti-depression targets of Kaixin San(KXS)in the brain tissue with"target fishing"strategy,and explored the target-associated pharmacological signaling pathways to reveal the anti-depression molecular mechanism of KXS.The Balb/c mouse model of depression was established by chronic unpredictable mild stress(CUMS)and the anti-depression effect of KXS was evaluated by forced swimming test and sucrose preference test.KXS active components were bonded to the benzophenone-modified magnetic nanoparticles by photocrosslinking reaction for capturing target proteins from cortex,thalamus and hippocampus of depressive mice.The target proteins were identified by liquid chromatography-mass spectrometry/mass spectrometry(LC-MS/MS).The enrichment analysis on signaling pathways was performed by Cytoscape.The potential biological functions of targets were verified by immunohistochemistry and Western blot assay.The results showed that KXS significantly improved the behavioral indexes.There were 64,91,and 44 potential targets of KXS identified in cortex,thalamus,and hippocampus,respectively,according to the target identification experiment.The functions of these targets were mainly associated with vasopressin-regulated water reabsorption,salmonella infection,thyroid hormone synthesis,and other signaling pathways.Besides,the results of immunohistochemistry and Western blot showed that KXS up-regulated the expressions of argipressine(AVP)in the cortex,heat shock protein 60(HSP60),cytochrome C oxidase 4(COX4),and thyrotropin-releasing hormone(TRH)in the thalamus,and down-regulated the expressions of tumor necrosis factor-α(TNF-α)and nuclear factor kappa B(NF-κB)p65 in the thalamus.Therefore,KXS may exert anti-depression effect through regulating vasopressin signaling pathway in the cortex and inflammation,energy metabolism,and thyroid hormone signaling pathways in the thalamus,and the effect of KXS on hippocampus is not significant.

关 键 词：抑郁 开心散 靶点鉴定 神经炎症 能量代谢 甲状腺激素 

分 类 号：R285.5[医药卫生—中药学]