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|a Available evidence suggests the involvement of microRNAs (miRNAs) in the pathological process of several diseases. Nonetheless, molecular mechanism underlying biological effects of miRNAs such as pacemaker exosome-derived miR-127-5p in embryonic-like stem cells (ESCs) differentiation into pacemaker cell is yet to be clarified. Through real time quantitative polymerase chain reaction (qPCR) or western blotting (WB) techniques, levels of miRNAs, miR-127-5p, and NKx2.5 expressions were quantitatively measured. Cellular differentiation (CD) was probed with flow cytometric (FC) and WB techniques. Prediction of miR-127-5p association with NKx2.5 was studied through bioinformatics tools before verification with luciferase assays. Promotion of ESCs differentiation to pacemaker through miR-127-5p was measured with qPCR and WB techniques. Through the same assaying methods, up-regulation of pace-making genes (Shox2, HCN4, Cx45, Tbx3 and Tbx18) expression was observed in Nkx2.5 knockdown group. However, Nkx2.5 expression was down-regulated during differentiation of pacemaker-like cells into ESCs. Furthermore, techniques (such as qPCR, WB, immunofluorescent staining and FC) were employed to demonstrate that overexpressed miR-127-5p could suppress NKx2.5 expression. Through NKx2.5 targeting, ESCs could be differentiated into pacemaker-like cells with miR-127-5p possibly serving as a crucial positive regulator. On the account of our findings, further researches are needed to unearth the possible underlying mechanism and role of exosome-derived miRNAs in cell signaling. © 2022 Elsevier Inc.
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