摘要
该研究主要是探讨六味地黄丸对卵巢早衰小鼠卵巢及皮质区雌性生殖干细胞的激活并促进其增殖作用进而改善卵巢功能。将ICR小鼠随机分为对照组、模型组、六味地黄丸组、六味地黄丸加倍组和戊酸雌二醇组。采用环磷酰胺腹腔注射制备卵巢早衰小鼠模型,造模成功后,连续灌胃六味地黄丸或者戊酸雌二醇治疗28 d并制备阴道涂片观察动情周期并记录体质量,末次给药后处死、取材,酶联免疫吸附测定(ELISA)检测血清雌二醇(E_(2))、促卵泡激素(FSH)、促黄体激素(LH)、抗缪勒管激素(AMH)水平;苏木素-伊红(HE)染色观察卵巢形态并计数各级卵泡,以评估卵巢功能;免疫组织化学检测卵巢雌性生殖干细胞标志物vasa基因同源类似物(MVH)表达及免疫荧光染色法采用MVH和增殖细胞核抗原(PCNA)共标检测雌性干细胞特异性标志物表达及定位;Western blot检测小鼠卵巢中MVH、八聚体结合转录因子4(Oct4)、PCNA蛋白表达。实验结果显示,与对照组相比,模型组小鼠动情周期紊乱,卵巢指数下降,闭锁卵泡增多,其他各级卵泡减少,FSH和LH水平明显上调,而AMH和E_(2)出现明显下降,显示造模成功。给予六味地黄丸、戊酸雌二醇治疗后,改善了激素水平、闭锁卵泡数量减少、其他各级卵泡数量增多,卵巢组织MVH标志蛋白、PCNA增殖蛋白增加。上述结果显示六味地黄丸调整卵巢早衰小鼠动情周期及激素紊乱,促进雌性生殖干细胞增殖,改善卵巢早衰卵泡发育,进而改善卵巢功能。
This study primarily investigates the effect of Liuwei Dihuang Pills on the activation and proliferation of female germline stem cells(FGSCs)in the ovaries and cortex of mice with premature ovarian failure(POF),and how it improves ovarian function.ICR mice were randomly divided into the control group,model group,Liuwei Dihuang Pills group,Liuwei Dihuang Pills double-dose group,and estradiol valerate group.A mouse model of POF was established by intraperitoneal injection of cyclophosphamide.After successful modeling,the mice were treated with Liuwei Dihuang Pills or estradiol valerate for 28 days.Vaginal smears were prepared to observe the estrous cycle and body weight.After the last administration,mice were sacrificed and sampled.Serum levels of estradiol(E_(2)),follicle-stimulating hormone(FSH),luteinizing hormone(LH),and anti-Müllerian hormone(AMH)were measured by enzyme-linked immunosorbent assay(ELISA).Hematoxylin-eosin(HE)staining was used to observe ovarian morphology and to count follicles at all stages to evaluate ovarian function.Immunohistochemistry was used to detect the expression of mouse vasa homolog(MVH),a marker of ovarian FGSCs.Immunofluorescence staining,using co-labeling of MVH and proliferating cell nuclear antigen(PCNA),was used to detect the expression and localization of specific markers of FGSCs.Western blot was employed to assess the protein expression of MVH,octamer-binding transcription factor 4(Oct4),and PCNA in the ovaries.The results showed that compared with the control group,the model group exhibited disordered estrous cycles,decreased ovarian index,increased atretic follicles,and a reduced number of follicles at all stages.FSH and LH levels were significantly elevated,while AMH and E_(2) levels were significantly reduced,indicating the success of the model.After treatment with Liuwei Dihuang Pills or estradiol valerate,hormone levels improved,the number of atretic follicles decreased,and the number of follicles at all stages increased.MVH marker protein and PCNA proliferative protein expression in ovarian tissue also increased.These results suggest that Liuwei Dihuang Pills regulate estrous cycles and hormone disorders in POF mice,promote the proliferation of FGSCs,improve follicular development in POF mice,and enhance ovarian function.
作者
姜波
张文燕
林广娣
马晓晴
兰国霞
仲佳雯
秦岭
买佳丽
李晓荣
JIANG Bo;ZHANG Wen-yan;LIN Guang-di;MA Xiao-qing;LAN Guo-xia;ZHONG Jia-wen;QIN Ling;MAI Jia-li;LI Xiao-rong(College of Traditional Chinese Medicine,Ningxia Medical University,Yinchuan 750004,China;General Hospital of Ningxia Medical University,Yinchuan 750000,China)
出处
《中国中药杂志》
北大核心
2025年第9期2495-2504,共10页
China Journal of Chinese Materia Medica
基金
国家自然科学基金项目(82260947)
宁夏自然科学基金项目(2023AAC05059)
2024年大学生创新创业训练计划项目(校级)(X202410752110)。
关键词
六味地黄丸
卵巢早衰
雌性生殖干细胞
ICR小鼠
Liuwei Dihuang Pills
premature ovarian failure
female germline stem cells
ICR mice
