Abstract
This study aims to elucidate the mechanisms by which microRNA-143-5p (miR-143-5p) targets runt-related transcription factor 2 (Runx2) in the differentiation of dental pulp stem cells (DPSCs) into odontoblasts, through regulating the osteoprotegerin receptor activator of the nuclear factor-κB ligand (OPG/RANKL) signaling pathway. Following transfection, DPSCs were divided into blank, control, miR-143-5p mimics, miR-143-5p inhibitors, miR-143-5p inhibitors + siRunx2 and siRunx2 groups. Alkaline phosphatase (ALP) activity and mineralized nodules were detected using ALP kit and alizarin red staining. Quantitative reverse transcriptase real time PCR (qRT-PCR) was conducted to measure mRNA expressions of miR-143-5p, Runx2, OPG, and RANKL. Western blotting was used to assess protein expression of odontoblast differentiation-related proteins. Transwell assay and an extracellular matrix (ECM) adhesion cell assay were employed to examine cell migration and cell adhesion. Compared with the blank group, the miR-143-5p mimics and siRunx2 groups showed decreased ALP activity, decreased mineralized nodules and displays of calcium. Fewer migrated cells, weakened cell adhesion, decreased protein expression of dentin phosphoprotein (DPP), dentin sialoprotein (DSP), dentin matrix protein 1 (DMP1), osteopontin (OPN), bone sialoprotein (BSP), osteocalcin (OCN), OPG and Runx2, and increased RANKL protein expressions were observed. Additionally, opposite results were observed in the miR-143-5p inhibitors group, demonstrating that down-regulated miR-143-5p promotes the differentiation of DPSCs into odontoblasts by enhancing Runx2 expression via the OPG/RANKL signaling pathway. Based on findings in this study, it is postulated that the enhancement of Runx2 expression via the regulation of the OPG/RANKL signaling pathway could be a beneficial approach for dental pulp regeneration. J. Cell. Biochem. 119: 536-546, 2018. © 2017 Wiley Periodicals, Inc.
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