人参皂苷Rh2通过mTOR/ROS信号通路抑制卵巢颗粒细胞衰老  

Ginsenoside Rh2 inhibits ovarian granulosa cell senescence through mTOR/ROS signaling pathway

作　　者：张杨[1] 姚美玉[1] 谷玥儒 孙淼[1] Zhang Yang;Yao Meiyu;Gu Yueru;Sun Miao(2nd Department of Gynecology,the First Affiliated Hospital of Heilongjiang Chinese Medicine University,Harbin 150036,China)

机构地区：[1]黑龙江中医药大学附属第一医院妇科二科,黑龙江哈尔滨150036

出　　处：《实用药物与临床》2024年第11期801-808,共8页Practical Pharmacy and Clinical Remedies

基　　金：中国博士后科学基金(2019M661320);国家自然科学基金资助项目(81804138、82174195);黑龙江省中医药科研项目(ZHY2020‑101)。

摘　　要：目的探讨人参及其主要活性成分人参皂苷Rh2抗胰岛素抵抗导致的卵巢颗粒细胞衰老的作用及机制。方法采用网络药理学、分子对接和细胞热位移分析法(Cellular thermal shift assay,CETSA)检测人参抗卵巢储备减少(Diminished ovarian reserve,DOR)和衰老的核心分子及其潜在的下游信号通路。用胰岛素(0.5μg/ml,72 h)处理人卵巢颗粒细胞(KGN)作为卵巢衰老的体外模型,将KGN细胞分为3组:对照组、胰岛素组(0.5μg/ml,72 h)以及胰岛素(0.5μg/ml,72 h)+人参皂苷Rh2组(2μmol/L,24 h)。β-半乳糖苷酶染色、CCK-8、活性氧检测试剂盒、免疫蛋白印迹以及qRT-PCR检测KGN细胞活性以及衰老相关因子(p53、p21)的表达水平。结果网络药理学筛选出人参有效成分、DOR和衰老核心度最高的10个共同靶点,即TNF、ESR1、MAPK3、PPARG、STAT3、HSP90AA1、NR3C1、mTOR、AR以及NOS3。人参中主要的活性成分为人参皂苷Rh2。与胰岛素组相比,人参皂苷Rh2(2μmol/L,24 h)能够降低KGN细胞中衰老细胞占比及活性氧(ROS)活性;同时,人参皂苷Rh2能够抑制KGN细胞中p53以及p21的表达升高。分子对接结果显示,mTOR是人参皂苷Rh2抗DOR以及衰老的关键靶点。过表达mTOR能够逆转人参皂苷Rh2抑制KGN细胞的活性降低和细胞衰老,逆转人参皂苷Rh2抑制KGN细胞ROS水平下降。结论人参皂苷Rh2可能通过调控mTOR/ROS抑制胰岛素抵抗导致的卵巢颗粒细胞衰老,从而发挥抵抗DOR的作用。Objective To investigate the effect and mechanism of ginsenoside Rh2 on ovarian granulosa cell aging induced by insulin resistance.Methods Network pharmacology,molecular docking,and Cellular Thermal Shift Assay(CETSA)were adopted to identify core molecules in ginseng that targeted diminished ovarian reserve(DOR)and senescence,along with potential downstream signaling pathways.Use insulin(0.5μg/ml,72 h)to induce aging in human ovarian granulosa cells(KGN),which served as an in vitro model for ovarian aging.The KGN cells were divided into three groups:control,insulin-treated(0.5μg/ml,72 h),and insulin-treated(0.5μg/ml,72 h)+Rh2 ginsenoside(2μmol/L,24 h)groups.Theβ-galactosidase staining,CCK-8 assay,reactive oxygen species assay,immunoblotting,and qRT-PCR were performed to assess KGN cell viability and the expression levels of aging-related factors(p53 and p21).Results The results of network pharmacology revealed 10 common targets with the highest core degree in active component in ginseng,DOR and aging:TNF,ESR1,MAPK3,PPARG,STAT3,HSP90AA1,NR3C1,mTOR,AR and NOS3.Ginsenoside Rh2(2μmol/L,24 h),the main active component in ginseng,could decrease the proportion of senescence and ROS activity in KGN cells.Meanwhile,ginsenoside Rh2 could inhibit the increased expression of p53 and p21 in KGN cells.Molecular docking results indicated that mTOR was a key target for the anti-DOR and senescence of ginsenoside Rh2.Overexpression of mTOR could reverse the decreased viability and senescence of KGN cells by ginsenoside Rh2,and reverse the decreased ROS level of KGN cells by ginsenoside Rh2.Conclusion Ginsenoside Rh2 can inhibit insulin-induced ovarian granulosa cell senescence by regulating mTOR/ROS,thus playing an anti-DOR role.

关 键 词：人参皂苷RH2 胰岛素抵抗 卵巢储备减少 MTOR 

分 类 号：R285[医药卫生—中药学]