Abstract
Abstract Study question Whether other forms of intercellular communication such as extracellular vesicles (EVs) participate in stem cell maintenance in the human endometrium. Summary answer The present study revealed the involvement of EVs as a novel intercellular communication method between endometrial mesenchymal stem cells (eMSC) and their surrounding niche. What is known already In human endometrium, many studies have demonstrated the importance of EVs in mediating various physiological as well as pathological processes. Our group has demonstrated that the myometrial cells are candidate niche cells of eMSC modulating their biological function. We also demonstrate the Notch signaling pathway is involved in endometrial stem cell regulation. Although classical Notch signaling relies on direct cell contact dependent interactions, this pathway can also be activated at a distance by EVs containing Notch ligands. Therefore, we hypothesized that certain Notch ligand(s) are packaged into the myometrial EVs to mediate stem cell functions. Study design, size, duration Sequential beading with magnetic beads coated with anti-CD140b and anti-CD146 antibodies was used to isolate eMSC (CD140b+CD146+ cells) from endometrial tissues from proliferative (n = 15) and secretory (n = 10) phase. Myometrial derived EVs (n = 17) were isolated by ultracentrifugation. The EVs were characterized by transmission electron microscopy and western blotting. Participants/materials, setting, methods Endometrial samples obtained from women aged 41 – 52 years undergoing total abdominal hysterectomy. EMSC were cocultured with myometrial EVs and the percentage of eMSC was analysed by flow cytometry. Blockage the secretion of EVs was performed by transfection of Rab27a siRNA. Western blot analysis and gene silencing approach validated the role of Notch signaling in eMSC. The therapeutic features of transplanted eMSC/myometrial EVs was determined using the mouse injured endometrial model. Main results and the role of chance EVs released from myometrial cells can be internalized by eMSC, leading to a significant effect on stemness of eMSC. Pharmacological inhibition of Notch signaling with DAPT or silencing of Notch 1 nullified these stimulatory effects. Myometrial EVs contains a high amount of the notch ligand – JAG1, thus inducing a strong Notch activity in eMSC. When JAG1 was silenced in the myometrial EVs, the self-renewal and clonogenic activity was reduced. Combined transplantation of eMSC with myometrial EVs improves the therapeutic effect of eMSC in endometrial regeneration in vivo. The observed therapeutic feature was potentially achieved by elevating the cell proliferation and suppressing apoptosis in the injured mouse endometrium. Limitations, reasons for caution Whether other signaling pathway are involved in the stimulatory effect of myometrial EVs in eMSC activities remains unknown. Also, EVs derived from other endometrial cell types i.e. epithelial cells may also regulate eMSC function. Wider implications of the findings Our findings revealed that myometrial EVs stimulated the biological function of eMSC via the Notch signaling. In the near future, it would be important to explore whether Notch signaling pathway plays a role in the injured mouse endometrium transplanted with eMSC/myometrial EVs. Trial registration number Not applicable
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