摘要
以二氧化乙烯基环己烯(VCD)联合高脂饲料建立围绝经期小鼠模型,评价人参固本口服液联合荆防颗粒对卵巢衰老及代谢功能的改善作用,并结合网络药理学与代谢组学探讨其机制。该实验将动物分为正常组,模型组,人参固本口服液低(3.6 mL·kg^(-1))、中(7.2 mL·kg^(-1))、高剂量(14.4 mL·kg^(-1))组,荆防颗粒组(4 g·kg^(-1)),联合用药组(人参固本口服液14.4 mL·kg^(-1)+荆防颗粒4 g·kg^(-1)),阳性药组(戊酸雌二醇0.18 mg·kg^(-1))。全自动生化分析仪检测血清中总胆固醇(TC)、甘油三酯(TG)、高密度脂蛋白胆固醇(HDL-C)、低密度脂蛋白胆固醇(LDL-C)的水平;酶联免疫吸附测定法(ELISA)检测血清中雌二醇(E_(2))、促卵泡生成素(FSH)、促黄体生成素(LH)、抗苗勒管激素(AMH)的水平;病理组织切片苏木素-伊红(HE)染色;利用网络药理学预测潜在作用靶点及通路,非靶向代谢组学检测血清代谢物变化,并采用Western blot检测卵巢雌激素受体(ERα),磷脂酰肌醇3-激酶(PI3K)、蛋白激酶B(Akt)、哺乳动物雷帕霉素靶蛋白(mTOR)、叉头框蛋白O1(FoxO1)及其磷酸化蛋白表达。结果显示,与模型组相比,联合用药可显著抑制体质量增长、降低Lee′s指数,改善TC、HDL-C、LDL-C及E_(2)、FSH、LH、AMH紊乱;代谢组学提示模型组磷脂及支链氨基酸代谢异常,主要富集于泛酸和辅酶A生物合成,缬氨酸、亮氨酸、异亮氨酸生物合成通路等;网络药理学筛选出雌激素受体1(ESR1)、丝裂原活化蛋白激酶1(MAPK1)、过氧化物酶体增殖物激活受体γ(PPARG)、白细胞介素6(IL6)等核心靶点,涉及雌激素、PI3K/Akt、FoxO信号通路;Western blot结果表明,联合用药上调ERα、磷酸化PI3K(p-PI3K)/PI3K、磷酸化Akt(p-Akt)/Akt、磷酸化mTOR(p-mTOR)/mTOR及磷酸化FoxO1(p-FoxO1)/FoxO1蛋白表达水平,改善卵巢能量代谢与凋亡失衡,从而发挥卵巢功能保护作用。综上,人参固本口服液联合荆防颗粒可多靶点调控卵巢代谢与信号通路,改善围绝经期小鼠卵巢功能,为中医药干预卵巢早衰及围绝经期综合征提供实验依据。
A perimenopausal mouse model was established using 4-vinylcyclohexene diepoxide(VCD)combined with a high-fat diet to investigate the regulatory effects of Renshen Guben Oral Liquid in combination with Jingfang Granules on ovarian aging and metabolic function,integrating with metabolomics and network pharmacology to elucidate the underlying mechanisms.Animals were allocated into normal,model,low-(3.6 mL·kg^(-1)),medium-(7.2 mL·kg^(-1)),and high-dose(14.4 mL·kg^(-1))Renshen Guben Oral Liquid,Jingfang Granules,combination treatment(14.4 mL·kg^(-1) Renshen Guben Oral Liquid+4 g·kg^(-1) Jingfang Granules),and positive control(estradiol valerate,0.18 mg·kg^(-1))groups.Serum levels of total cholesterol(TC),triglyceride(TG),high-density lipoprotein cholesterol(HDL-C),and low-density lipoprotein cholesterol(LDL-C)were measured using an automatic biochemical analyzer.Serum estradiol(E_(2)),follicle-stimulating hormone(FSH),luteinizing hormone(LH),and anti-Müllerian hormone(AMH)were determined by enzyme-linked immunosorbent assay(ELISA).Ovarian histopathology was examined by hematoxylin-eosin(HE)staining.Potential targets and pathways were predicted through network pharmacology,and non-targeted metabolomics was applied to detect serum metabolite alterations.Western blot was employed to detect ovarian protein expression of estrogen receptor alpha(ERα),phosphatidylinositol 3-kinase(PI3K),protein kinase B(Akt),mammalian target of rapamycin(mTOR),forkhead box protein O1(FoxO1),and their phosphorylated forms.The results showed that compared to the model group,combination treatment markedly attenuated body weight gain,reduced Lee′s index,restored the levels of TC,HDL-C,and LDL-C,and alleviated the disorders in E_(2),FSH,LH,and AMH.Metabolomic analysis revealed disturbances in phospholipid and branched-chain amino acid metabolism in the model group,with enrichment in pantothenate and CoA biosynthesis as well as valine,leucine,and isoleucine biosynthesis pathways.Network pharmacology identified core targets including estrogen receptor 1(ESR1),mitogen-activated protein kinase 1(MAPK1),peroxisome proliferator-activated receptor gamma(PPARG),and interleukin 6(IL6),which were associated with estrogen,PI3K/Akt,and FoxO signaling pathways.Western blot further demonstrated that combination treatment upregulated the protein expression of ERα,p-PI3K/PI3K,p-Akt/Akt,p-mTOR/mTOR,and p-FoxO1/FoxO1,thereby improving ovarian energy metabolism and apoptotic imbalance,ultimately exerting protective effects on ovarian function.These findings suggested that the synergistic use of Renshen Guben Oral Liquid and Jingfang Granules exerts multi-targeted regulation of ovarian metabolism and signaling pathways and improves ovarian function in perimenopausal mice,providing experimental evidence for TCM intervention in premature ovarian failure and perimenopausal syndrome.
作者
巩立媛
张萌迪
赵文歆
姚方方
杜建才
姚景春
王恩力
张贵民
GONG Li-yuan;ZHANG Meng-di;ZHAO Wen-xin;YAO Fang-fang;DU Jian-cai;YAO Jing-chun;WANG En-li;ZHANG Gui-min(Graduate School,Tianjin University of Traditional Chinese Medicine,Tianjin 301617,China;State Key Laboratory of Integration and Innovation of Classic Formula and Modern Chinese Medicine,Lunan Pharmaceutical Group Co.,Ltd.,Linyi 273400,China)
出处
《中国中药杂志》
北大核心
2026年第4期1031-1042,共12页
China Journal of Chinese Materia Medica
基金
山东省重点研发计划项目(2021CXGC010508)。
