基于AGE-RAGE信号通路探讨肾气丸治疗肾虚排卵障碍的作用机制  

Mechanism of Shenqi Pills(肾气丸)in Treatment of Ovulation Disorder with Shenxuzheng(肾虚证)Based on AGE-RAGE Signaling Pathways

作　　者：刘毓 孙理军[1] 佟雅婧 LIU Yu;SUN Lijun;TONG Yajing(Shaanxi University of Chinese Medicine,Xianyang 712046)

机构地区：[1]陕西中医药大学,咸阳712046

出　　处：《中药药理与临床》2024年第11期21-28,共8页Pharmacology and Clinics of Chinese Materia Medica

基　　金：陕西省教育厅重点实验室项目(编号:19JS014);陕西省教育厅重点研究项目(编号:SMZX202102)。

摘　　要：目的:通过网络药理学探究肾气丸改善排卵障碍(Ovulation disorder,OD)的作用机制,并通过动物体内实验验证分析。方法:通过网络药理学筛选活性成分与相关靶点,构建“中药-活性成分-靶点”作用网络及PPI蛋白互作网络,用R语言对交集靶点进行GO和KEGG分析。根据网络药理学结果进行动物实验验证,采用双下肢肌注氢化可的松注射液诱导模型建立,造模成功大鼠给予肾气丸干预14 d后,阴道涂片观察性周期;HE染色观察卵巢组织病理变化;TUNEL染色检测大鼠卵巢颗粒细胞凋亡情况;检测血清FSH、LH、E_(2)、P、IL-1β、IL-6、TNF-α含量;Western-blot法检测卵巢组织AGEs、RAGE、ERK1/2、C-JUN蛋白表达。结果:确定肾气丸治疗OD的活性成分117个,治疗靶点93个,槲皮素、薯蓣皂苷元、谷甾醇、(+)-表儿茶素和二氢槲皮素可能为干预OD的关键成分;PPI网络筛选得到20个核心靶点;GO富集分析显示:分子功能中排名前3的分别是DNA结合转录因子结合、RNA聚合酶Ⅱ特异性DNA结合转录因子结合和DNA结合转录激活剂活性,生物过程排名前3的分别是对外来生物刺激的反应、上皮细胞增殖和氧化应激反应,细胞成分排名前3的是膜筏、膜微区和囊泡腔;KEGG分析提示AGE-RAGE信号通路在肾气丸治疗OD过程中起着关键作用。动物实验表明,肾气丸干预后可改善大鼠性周期紊乱;增加各级卵泡数量;减少卵泡闭锁;抑制卵巢颗粒细胞凋亡;使FSH、LH、E_(2)、P激素水平趋于正常;降低炎症反应;下调AGEs、RAGE、ERK、C-JUN蛋白表达,调控AGE-RAGE信号通路以改善OD。结论:肾气丸可能通过调节AGE-RAGE信号通路上AGEs、RAGE、ERK、C-JUN蛋白表达以改善激素水平、降低炎症反应、抑制颗粒细胞凋亡,从而达到改善OD的目的。Objective:To study the mechanism of Shenqi Pills(肾气丸)in improving ovulation disorder(OD)through network pharmacology and to validate the findings through animal experiments.Methods:Active components and related targets were screened by network pharmacology,and a"drug-active component-target"network and a protein-protein interaction(PPI)network was constructed.Gene Ontology(GO)and Kyoto Encyclopedia of Genes and Genomes(KEGG)analyses were performed on the intersecting targets using R language.According to the results of network pharmacology,animal experiments were conducted.A model was established by intramuscular injection of hydrocortisone in both hind limbs.After successful modeling,Shenqi Pills were administered for 14 days.Vaginal smears were used to observe estrous cycles.HE staining was used to observe histopathological changes in ovarian tissue.TUNEL staining was used to detect apoptosis in ovarian granulosa cells.Serum levels of FSH,LH,E_(2),P,IL-1β,IL-6,and TNF-α were measured to assess hormone levels and inflammatory response.Western blot was used to detect the expression of AGEs,RAGE,ERK1/2,and C-JUN proteins in ovarian tissue.Results:Shenqi Pills were found to contain 117 active components and 93 therapeutic targets for treating OD.Quercetin,diosgenin,β-sitosterol,(+)-epicatechin,and dihydroquercetin were identified as potential key components for OD intervention.The PPI network identified 20 core targets.GO enrichment analysis showed the top three molecular functions were DNA-binding transcription factor binding,RNA polymerase Ⅱ-specific DNA-binding transcription factor binding,and DNA-binding transcription activator activity.The top three biological processes were response to external biotic stimulus,epithelial cell proliferation,and oxidative stress response.The top three cellular components were membrane raft,membrane microdomain,and vesicle lumen.KEGG analysis indicated that the AGE-RAGE signaling pathway played a crucial role in the treatment of OD with Shenqi Pills.Animal experiments

关 键 词：肾气丸 网络药理学 肾虚 排卵障碍 晚期糖基化终末产物/糖基化终产物受体信号通路 

分 类 号：R73[医药卫生—肿瘤]