摘要
目的基于转录组学探讨疏肝补肾法治疗卵巢早衰(premature ovarian failure,POF)大鼠的作用机制。方法将30只SD大鼠采用随机数字表法分为空白组(n=10)、模型组(n=10)和治疗组(n=10),空白组未施加干预措施,模型组和治疗组采用慢性不可预知温和刺激法联合环磷酰胺化学诱导法建立肝郁肾虚POF大鼠模型并评价。成模后将大鼠随机平均分为模型组和治疗组,根据体表面积等效剂量折算原理,治疗组予以疏肝补肾方13.55 g/(kg·d)灌胃,空白组及模型组予以等剂量生理盐水灌胃,干预周期为3周。于造模前与给药干预后监测各组大鼠体质量变化,采用苏木精-伊红(HE)染色法观察卵巢组织形态学改变,并通过酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)检测血清卵泡刺激素(follicle-stimulating hormone,FSH)及雌二醇(estradiol,E_(2))水平,并采用RNA测序(RNA sequencing,RNA-seq)筛选差异表达基因(differentially expressed genes,DEGs)。结果造模结束后,与空白组比较,模型组大鼠体质量明显降低(P<0.01),血清FSH含量明显升高(P<0.01),血清E 2含量明显降低(P<0.01);提示造模成功。大鼠卵巢组织形态学显示,与空白组比较,模型组卵巢组织体积缩小,内部结构破坏,未成熟的闭锁卵泡数量增加,成熟卵泡较少;与模型组比较,治疗组卵巢组织体积增大,闭锁卵泡数量减少,可见较多各级卵泡。与模型组比较,治疗组卵巢指数增加(P<0.05),血清FSH降低(P<0.01),血清E 2升高(P<0.05)。空白组与模型组的转录组基因本体论(gene ontology,GO)富集分析显示,DEGs主要富集于慢性应激与氧化应激调控等相关通路。模型组与治疗组的转录组分析共鉴定出81个DEGs[筛选条件:错误发现率(false discovery rate,FDR)<0.005且差异倍数(|Fold Change|)>1.5],GO功能富集分析表明DEGs主要富集于氧化还原功能通路。其中,模型组存在显著上调的致癌基因(Tk1、Kif20b、Micall2、Enpp3、MAFK)。结论疏肝补肾法能有效调节POF大鼠性激素水平,并可能通过氧化还原途径,抑制致癌基因表达,从而改善卵泡发育环境。
Objective To explore the mechanism of dispersing stagnated liver qi and tonifying kidney method in treating premature ovarian failure(POF)rats based on transcriptomics.Methods Thirty SD rats were randomly divided into the blank group(n=10),the model group(n=10)and the treatment group(n=10)using a random number table method.The control group received no intervention.The model group and the treatment group were established using a chronic unpredictable mild stress protocol combined with cyclophosphamide-induced chemotherapy to create liver-depression and kidney-deficiency POF rat model.After model establishment,rats were randomly and equally assigned to the model and treatment groups.According to the principle of equivalent dose conversion based on body surface area,the treatment group was given 13.55 g/(kg·d)by gavage with the Shugan Bushen formula,while the blank group and the model group were given an equal dose of physiological saline by gavage.The intervention period were three weeks.Monitoring changes in body weight in each group of rats before modeling and after drug intervention.Hematoxylin eosin(HE)staining was used to observe the morphological changes of ovarian tissue,and enzyme-linked immunosorbent assay(ELISA)was used to detect the serum levels of follicle-stimulating hormone(FSH)and estradiol(E_(2)).RNA sequencing(RNA-seq)was used to screen differentially expressed genes(DEGs).Results After modeling,compared with the blank group,the body weight of the model group rats decreased significantly(P<0.01).The serum level of FSH was significantly increased(P<0.01),and the serum level of E 2 decreased significantly(P<0.01).The above suggesting that modelling was successful.Morphology of rats ovarian tissue showed that compared with the blank group,the model group had reduced ovarian tissue volume and internal structural damage,and an increase in the number of immature blocked follicles and fewer mature follicles.Compared with the model group,the treatment group had a larger volume of ovarian tissue,a reduced number of blocked follicles,and more follicles at all levels were visible.Compared with the model group,the ovarian index in the treatment group was increased(P<0.05),the serum level of FSH was decreased(P<0.01),and the level of E 2 was increased(P<0.05).Gene ontology(GO)enrichment analysis of transcriptome genes between the blank group and model group showed that DEGs were mainly enriched in pathways related to chronic stress and oxidative stress regulation.The transcriptome analysis of the model group and treatment group identified a total of 81 DEGs[screening conditions:false discovery rate(FDR)<0.005 and|fold change|>1.5].GO functional enrichment analysis showed that DEGs were mainly enriched in the redox functional pathway.Among them,there were significantly upregulated oncogenes(Tk1,Kif20b,Micall2,Enpp3,MAFK)in the model group.Conclusion The method of dispersing stagnated liver qi and tonifying kidney could effectively regulate the levels of sex hormones in POF rats,and may improve the follicular development environment by inhibiting the expression of oncogenes through the redox pathway.
作者
陈雯婕
练正涛
王洋
CHEN Wenjie;LIAN Zhengtao;WANG Yang(Traditional Chinese Medicine Syndrome Research Base,Fujian University of Traditional Chinese Medicine,Fuzhou 350122,China;School of Traditional Chinese Medicine,Fujian University of Traditional Chinese Medicine,Fuzhou 350122,China)
出处
《中西医结合研究》
2026年第1期33-39,共7页
Research of Integrated Traditional Chinese and Western Medicine
基金
福建省自然科学基金项目(No.2025J01938)
福建中医药大学校管课题(No.X2023028)。
关键词
卵巢早衰
疏肝补肾
转录组学
氧化应激
premature ovarian failure
dispersing stagnated liver qi and tonifying kidney
transcriptomics
oxidative stress
