六味地黄丸对卵巢早衰小鼠激素水平、氧化应激、颗粒细胞自噬的影响及机制  

Effects and Mechanisms of Liuwei Dihuang Pills on Hormone Levels,Oxidative Stress,and Granulosa Cell Autophagy in Premature Ovarian Failure Mice

作　　者：仲佳雯 姜波 张文燕 李晓荣[1,3,4] 秦岭 高婷 ZHONG Jiawen;JIANG Bo;ZHANG Wenyan;LI Xiaorong;QIN Ling;GAO Ting(General Hospital of Ningxia Medical University,Yinchuan 750000,China;College of Traditional Chinese Medicine of Ningxia Medical University,Yinchuan 750000,China;Key Laboratory of Fertility Maintenance,Ningxia Medical University,Yinchuan 750000,China;Key Laboratory of Ethnic Medicine Modernization,Ministry of Education,Ningxia Medical University,Yinchuan 750000,China)

机构地区：[1]宁夏医科大学总医院,银川75000 [2]宁夏医科大学中医学院,银川750000 [3]宁夏医科大学生育力保持重点实验室,银川750000 [4]宁夏医科大学少数民族医药现代化教育部重点实验室,银川750000

出　　处：《世界中医药》2025年第4期574-582,592,共10页World Chinese Medicine

基　　金：国家自然科学基金项目(82260947);宁夏自然科学基金项目(2023AAC05059)。

摘　　要：目的:探讨六味地黄丸对卵巢早衰小鼠模型激素水平、氧化应激状态以及颗粒细胞自噬的影响,并深入解析其可能的作用机制。方法:腹腔注射环磷酰胺(CTX)建立卵巢早衰小鼠模型,造模成功后小鼠分为模型组、六味地黄丸低剂量组、六味地黄丸高剂量组、戊酸雌二醇组、通路抑制剂组(NVP-BEZ235)、通路抑制剂+六味地黄丸高剂量组,连续灌胃28 d。采用酶联免疫吸附测定(ELISA)方法对小鼠血清中的激素及氧化相关指标进行检测,包括抗米勒管激素(AMH)、雌二醇(E2)、黄体生成素(LH)、促卵泡激素(FSH)、超氧化物歧化酶(SOD)、谷胱甘肽过氧化物酶(GSH-Px)以及丙二醛(MDA)等指标。处死后称取双侧卵巢计算卵巢指数;苏木精-伊红(HE)染色评估卵巢组织的病理变化;免疫组织化学检测卵巢组织中微管相关蛋白1轻链3(LC3)含量以及透射电镜下自噬体计数评估线自噬水平;蛋白质免疫印迹法及qPCR检测自噬及通路相关蛋白和基因的表达。结果:与模型组比较:六味地黄丸高剂量组E2、AMH、SOD、GSH-Px显著升高(P<0.05),FSH、LH、MDA显著降低(P<0.05);卵巢指数显著提高(P<0.05);增加小鼠黄体数,减少闭锁卵泡数(P<0.05);自噬体计数均降低,线粒体自噬相关标志物LC3降低,而螯合体1(P62)、磷酸化磷脂酰肌醇3-激酶/总磷脂酰肌醇3-激酶(p-PI3K/PI3K)、磷酸化蛋白激酶B/总蛋白激酶B(p-AKT/AKT)显著增加(P<0.05)。结论:六味地黄丸保护卵巢免受环磷酰胺引起的卵巢损伤和氧化应激;六味地黄丸可能通过激活PI3K/AKT/mTOR抑制颗粒细胞过度自噬来改善卵巢功能。Objective:To investigate the effects of Liuwei Dihuang Pills(LWDH)on hormone levels,oxidative stress,and granulosa cell autophagy in a mouse model of premature ovarian failure(POF),and to explore its potential mechanisms.Methods:POF in mice was induced by intraperitoneal injection of cyclophosphamide(CTX).After successful modeling,the mice were divided into the model group,low-dose LWDH group,high-dose LWDH group,estradiol valerate group,pathway inhibitor group(NVP-BEZ235),and pathway inhibitor+high-dose LWDH group.Mice were administered by gavage for 28 days.Hormones and oxidation-related markers in serum were measured using enzyme-linked immunosorbent assay(ELISA),including anti-Müllerian hormone(AMH),estradiol(E 2),luteinizing hormone(LH),follicle-stimulating hormone(FSH),superoxide dismutase(SOD),glutathione peroxidase(GSH-Px),and malondialdehyde(MDA).After euthanizing the mice,the ovaries were weighed to calculate the ovarian index.Hematoxylin-eosin(HE)staining was used to assess pathological changes in ovarian tissues.Immunohistochemistry was used to detect the content of microtubule-associated protein light chain 3(LC3)in ovarian tissues,and autophagosome counts were assessed by transmission electron microscopy to evaluate autophagy.Western blot and qPCR were performed to detect the expression of autophagy-related and signaling pathway-related proteins and genes.Results:Compared to the model group,the high-dose LWDH group showed significant increases in E 2,AMH,SOD,and GSH-Px(P<0.05),and significant decreases in FSH,LH,and MDA(P<0.05).The ovarian index was significantly increased(P<0.05),the number of corpora lutea was increased,and the number of atretic follicles was reduced(P<0.05).Autophagosome counts were significantly decreased,and the autophagy marker LC3 was reduced,while sequestosome 1(P62),phosphorylated phosphatidylinositol 3-kinase/total phosphatidylinositol 3-kinase(p-PI3K/PI3K),and phosphorylated protein kinase B/total protein kinase B(p-AKT/AKT)were significantly increased(P<0.05).Conclusion

关 键 词：六味地黄丸 卵巢早衰 激素 氧化应激 颗粒细胞自噬 环磷酰胺 磷脂酰肌醇3-激酶/蛋白激酶B/哺乳动物雷帕霉素靶蛋白信号通路 小鼠 

分 类 号：R285.5[医药卫生—中药学]