Study On The Effect And Mechanism Of Exosomes Derived From Human Umbilical Cord Mesenchymal Stem Cells On The Repair Of Endometrial Injury

Objective:After treatment with exosomes derived from human umbilical cord mesenchymal stem cells(hUCMSC-Exos),the endometrium repair of mice with endometrial injury,as well as the proliferation,apoptosis,and angiogenesis of the endometrium cells were detected.To explore the effect of hUCMSC-Exos on the damaged endometrium in a mouse endometrial injury model and its underlying mechanism.Methods:1.Exosomes were extracted from human umbilical cord mesenchymal stem cells using an exosome extraction kit.Western blotting was used to detect the labeled proteins of the explants,and the morphology and size of the extracts were observed by transmission electron microscopy.2.A mouse endometrial injury model was established with 95% alcohol.Then after injecting Di R-labeled hUCMSC-Exos into the mice,the fluorescence signal of mouse uterus was observed at different time points with an in vivo imaging system.3.All female mice were randomly divided into control group,saline group,and exosome group.The saline group were treated with saline after model were made with alcohol;exosome group were treated with hUCMSC-Exos.The endometrial thickness was observed by HE staining experiment.4.To detect apoptosis,cell proliferation and vascular regeneration in the uterus of each group of mice.The expression of Ki67 and CD31 in mouse uterus was detected by immunohistochemistry.Endometrial cell apoptosis was detected by TUNEL assay.The expression levels of Bax,Bcl-2 and Cleaved Caspase-3 proteins in the uterine tissues of the above three groups were detected by Western Blotting.The expression of VEGF and IGF-1 m RNA was detected by q RT-PCR.To detect the expression of PTEN/Akt pathway,related proteins,the protein expression levels of PTEN,Akt and p Akt were detected by Western Blotting.Results:1.The isolated exosomes were observed by transmission electron microscopy to be spherical with diameters ranging from 20-300 nm.Western blotting results confirmed that the extracted exosomes expressed the exosome-specific markers CD63,PDCD6 IP,HSPA8 and TSG101 and did not express β-Actin.2.Partial fluorescent signals were detected in the damaged side of the uterus at 6 and 12 hours after hUCMSC-Exos injection,and high intensity fluorescent signals were detected at 24 hours after injection.3.The results of HE staining indicated that the endometrial epithelial cells of mice in the modeling group were significantly damaged,with a significant decrease in thickness and the number of glands.In contrast,the morphology and thickness of endometrial glandular epithelial cells in the exosome group were significantly improved,and the number of glands was also increased compared with that in the modeling group,and the differences were statistically significant(P<0.05).4.Compared with the saline group,apoptosis was significantly reduced and the expression of Ki67 and CD31 was significantly increased in the exosome group,with statistically significant differences(P < 0.05).hUCMSC-Exos treatment resulted in a significant decrease in the expression of Bax and Cleaved Caspase-3,an increase in the expression of Bcl-2,and the m RNA expression levels of VEGF and IGF-1 were significantly upregulated.After hUCMSC-Exos treatment,the phosphorylation level of Akt was significantly increased and the expression of PTEN was significantly decreased.Conclusions:Human umbilical cord MSC-derived exosomes can increase endometrial thickness,gland number,promote vascular regeneration,and improve damaged endometrium in a mouse model of endometrial injury,which may act by activating the PTEN/Akt pathway to regulate the expression of the Bcl-2 family and angiogenesis-related factors.