Circ6401, a novel circular RNA, is implicated in repair of the damaged endometrium by Wharton’s jelly-derived mesenchymal stem cells through regulation of the miR-29b-1-5p/RAP1B axis

Abstract

Background: Accumulating evidence indicates that mesenchymal stem cells (MSCs) exert tissue repair effects and therapeutic angiogenesis through their noncoding RNAs (ncRNAs). Our previous studies showed that MSCs derived from Wharton’s jelly (WJ-MSCs) can ameliorate damaged human endometrium by promoting angiogenesis. There is limited information on the functions and mechanism of ncRNAs in MSC-induced endometrial repair, and additional studies are needed for more insights. Methods: Here, WJ-MSCs were cocultured with or without endometrial stromal cells (ESCs) damaged by mifepristone (cocultured group versus non-cocultured group). TUNEL staining assays, EdU proliferation assays, flow cytometry apoptosis assays, and western blot assays were performed to observe the reparative effect of WJ-MSCs on damaged ESCs. Subsequently, circular RNA (circRNA) and microRNA microarrays were performed between the two groups. A subset of top upregulated circRNAs was validated by qRT-PCR. The functions of circ6401 (hsa_circ_0006401) in WJ-MSCs were investigated using lentivirus-mediated circRNA overexpression assays. The subcellular localization of circ6401 and miR-29b-1-5p in WJ-MSCs was identified by double RNA fluorescence in situ hybridization. Dual-luciferase reporter assays and western blot assays were performed to elucidate the regulatory mechanisms among circ6401, miR-29b-1-5p, and RAP1B. Results: WJ-MSCs significantly improved ESC proliferation and upregulated the expression of vascular angiogenesis markers. Circ6401 was upregulated in WJ-MSCs cocultured with damaged ESCs, while miR-29b-1-5p was significantly downregulated. Furthermore, circ6401 was found to bind to miR-29b-1-5p and prevent it from decreasing the level of RAP1B, a crucial protein involved in the VEGF signaling pathway, which promoted angiogenesis and stimulated the proliferation of ESCs. Conclusions: Our results showed the abundance and regulation profiles of ncRNAs of WJ-MSCs during repair of damaged ESCs and, for the first time, clarified the underlying mechanism by which circ6401 promotes endometrial repair by WJ-MSCs; thus, demonstrating that circ6401 may serve as a potential therapeutic target.