中国全科医学2025，Vol.28Issue(8)：962-972,11.DOI:10.12114/j.issn.1007-9572.2023.0768
外泌体调节复发性流产小鼠子宫内膜细胞上皮间充质转化的作用机制研究
Exosomes Regulate Epithelial Mesenchymal Transition of Endometrial Cells in Mice with Recurrent Spontaneous Abortion
摘要
Abstract
Background Recurrent spontaneous abortion(RSA)is a common reproductive disorder closely linked with the weakened epithelial-mesenchymal transition(EMT)in the endometrium during embryo implantation.Endometrium-derived exosomes and miRNAs they transport are linked with RSA via the involvement in cellular communication and EMT,although the specific mechanisms have not yet fully revealed.Objective Focusing on the function of exosomes in EMT,this study aims to validate the regulatory role of endometrium-derived exosomes and miRNA-221 they transport in EMT of endometrial cells in RSA mice.Methods From October 2021 to May 2022,a total of 24 eight-week-old female mice in the specific pathogen-free(SPF)level were randomly assigned into the control group(n=12)and RSA group(n=12)for modeling.After RSA modeling,male and female mice were housed together.On the 4th day of pregnancy,female mice were sacrificed for isolating endometrial epithelial cells.They were cultured for extracting exosomes,which were identified by transmission electron microscopy and Western blotting.Cell proliferation,invasion,and migration abilities were evaluated using CCK-8 assay,Transwell assay,and fluorescence-activated cell sorting(FACS)before and after exosome interventions.Expression levels of EMT-related genes[E-cadherin,N-cadherin,Vimentin,matrix metalloproteinase 7(MMP-7),matrix metalloproteinase 9(MMP-9)]and exosome marker proteins[CD63,CD81,tumor susceptibility gene 101(TSG101)]were measured.Expression levels of EMT marker proteins(CD24-APC,CD44-FITC)in exosome-treated cells were analyzed.Transfection of miRNA-221 mimics and inhibitors was performed to analyze the effects of exosome-derived miRNA-221 on EMT of endometrial epithelial cells in RSA mice.Relevant analyses were conducted using Western blot and dual-luciferase reporter assay.Results Abnormal EMT was observed in the endometrial epithelial cells of mice in RSA group,and the mouse endometrial epithelial cells were able to secrete exosomes.The proliferation rate,migration ability,invasion ability,and CD44+/CD24+ratio of cells in the RSA group treated with exosomes were significantly higher than those measured in cell supernatant of RSA group without exosome intervention(P<0.05).Compared with those in the cell supernatant of RSA group,induction of exosomes significantly downregulated EMT marker protein E-cadherin,but upregulated stromal marker proteins N-cadherin and Vimentin,as well as matrix marker proteins MMP-7 and MMP-9(P<0.05).The expression level of miRNA-221 in the RSA group was significantly lower than that of the control group(P<0.05).Compared with those transfected with miRNA negative control,transfection of miRNA-221 mimic significantly activated the phosphatidylinositol 3-kinase(PI3K)-AKT pathway,nuclear factor-kappa B(NF-κB)pathway,and p38 pathway(P<0.05),but downregulated mRNA level of PTEN(P<0.05).Transfection of miRNA-221 inhibitor significantly upregulated mRNA level of PTEN(P<0.05).Conclusion Endometrium-derived exosomal miRNA-221 negatively regulates the PTEN gene,activates the PI3K-AKT pathway,promotes EMT in endometrial epithelial cells,and influences the pregnant outcome of RSA mice.关键词
复发性流产/上皮间充质转化/外泌体/miRNA-221/Pten基因/PI3K-AKT通路Key words
Recurrent spontaneous abortion/Epithelial-mesenchymal transition/Exosomes/miRNA-221/PTEN gene/PI3K-AKT pathway分类
医药卫生引用本文复制引用
李梦元,何军琴,李冠杉,韩倩,李思瑶,郑舒畅,辛明蔚,尹晓丹,王景尚,武颖..外泌体调节复发性流产小鼠子宫内膜细胞上皮间充质转化的作用机制研究[J].中国全科医学,2025,28(8):962-972,11.基金项目
国家自然科学基金面上资助项目(82074477) （82074477）
北京市自然科学基金面上资助项目(7222268) （7222268）
