Magnesium-enriched microenvironment promotes odontogenic differentiation in human dental pulp stem cells by activating ERK/BMP2/Smads signaling

Yuanyuan Kong,Yingqun Zhong,Jianmao Zheng,Ke Xu,Xiaoli Hu,Buling Wu

doi:10.1186/s13287-019-1493-5

Abstract

BackgroundMagnesium (Mg2+)-enriched microenvironment promotes odontogenic differentiation in human dental pulp stem cells (DPSCs), but the regulatory mechanisms remain undefined. The aim of this work was to assess magnesium’s function in the above process and to explore the associated signaling pathway.MethodsDPSCs underwent culture in odontogenic medium with the addition of 0, 1, 5, or 10 mM MgCl2. Intracellular Mg2+ levels in DPSCs were evaluated flow cytometrically using Mag-Fluo-4-AM. Mg2+-entry was inhibited by TRPM7 inhibitor 2-aminoethoxydiphenyl borate (2-APB). RNA-Sequencing was carried out for assessing transcriptome alterations in DPSCs during odontogenic differentiation associated with high extracellular Mg2+. KEGG pathway analysis was performed to determine pathways related to the retrieved differentially expressed genes (DEGs). Immunoblot was performed for assessing magnesium’s role and exploring ERK/BMP2/Smads signaling.ResultsMg2+-enriched microenvironment promoted odontogenic differentiation in DPSCs via intracellular Mg2+ increase. Consistently, the positive effect of high extracellular Mg2+ on odontogenic differentiation in DPSCs was blocked by 2-APB, which reduced Mg2+ entry. RNA-sequencing identified 734 DEGs related to odontogenic differentiation in DPSCs in the presence of high extracellular Mg2+. These DEGs participated in many cascades such as MAPK and TGF-β pathways. Consistently, ERK and BMP2/Smads pathways were activated in DPSCs treated with high extracellular Mg2+. In agreement, ERK signaling inhibition by U0126 blunted the effect of high extracellular Mg2+ on mineralization and odontogenic differentiation in DPSCs. Interestingly, BMP2, BMPR1, and phosphorylated Smad1/5/9 were significantly decreased by U0126, indicating that BMP2/Smads acted as downstream of ERK.ConclusionsMg2+-enriched microenvironment promotes odontogenic differentiation in DPSCs by activating ERK/BMP2/Smads signaling via intracellular Mg2+ increase. This study revealed that Mg2+-enriched microenvironment could be used as a new strategy for dental pulp regeneration.

Highlights

Dental pulp regeneration may be a potential treatment for managing permanent teeth undergoing necrosis [1]
There was no difference between the 5 and 10 mM Mg2+ groups (Fig. 3). These results showed that the Mg2+-enriched microenvironment promoted the odontogenic differentiation of dental pulp stem cells (DPSCs)
The results showed that Bone morphogenetic protein 2 (BMP2), Bone morphogenetic protein receptor 1 (BMPR1), and phosphorylated Smad1/5/9 were significantly increased in DPSCs cultured in odontogenic medium containing high extracellular Mg2+(Fig. 6c)

Summary

Introduction

Dental pulp regeneration may be a potential treatment for managing permanent teeth undergoing necrosis [1]. Evidence indicates mutations of the Mg2+ transporters TRPM7 and CNNM4 result in deficient dentin mineralization, confirming Mg2+ involvement in tooth development [11, 12]. Our previous study demonstrates that the Mg2+ transporter Magt plays an important role in odontogenic differentiation of bone marrow MSCs by regulating intracellular Mg2+ [14]. Studies have found high extracellular Mg2+ and its transporter regulate odontogenic differentiation in human DPSCs, with involvement in dentin mineralization [4, 15]. Magnesium (Mg2+)-enriched microenvironment promotes odontogenic differentiation in human dental pulp stem cells (DPSCs), but the regulatory mechanisms remain undefined. The aim of this work was to assess magnesium’s function in the above process and to explore the associated signaling pathway

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Discussion

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