EphB4/ TNFR2/ERK/MAPK signaling pathway comprises a signaling axis to mediate the positive effect of TNF-\u03b1 on osteogenic differentiation

Yu Zhang,Shaohua Ge,Pishan Yang,Limei Wang,Jin Zhang,Chengzhe Yang

doi:10.1186/s12860-020-00273-2

Abstract

BackgroundLow concentrations of tumor necrosis factor-alpha (TNF-α) and its receptor TNFR2 are both reported to promote osteogenic differentiation of osteoblast precursor cells. Moreover, low concentrations of TNF-α up-regulate the expression of EphB4. However, the molecular mechanisms underlying TNF-α-induced osteogenic differentiation and the roles of TNFR2 and EphB4 have not been fully elucidated.ResultsThe ALP activity, as well as the mRNA and protein levels of RUNX2, BSP, EphB4 and TNFR2, was significantly elevated in MC3T3-E1 murine osteoblast precursor cells when stimulated with 0.5 ng/ml TNF-α. After TNFR2 was inhibited by gene knockdown with lentivirus-mediated shRNA interference or by a neutralizing antibody against TNFR2, the pro-osteogenic effect of TNF-α was partly reversed, while the up-regulation of EphB4 by TNF-α remained unchanged. With EphB4 forward signaling suppressed by a potent inhibitor of EphB4 auto-phosphorylation, NVP-BHG712, TNF-α-enhanced expressions of TNFR2, BSP and Runx2 were significantly decreased. Further investigation into the signaling pathways revealed that TNF-α significantly increased levels of p-JNK, p-ERK and p-p38. However, only the p-ERK level was significantly inhibited in TNFR2-knockdown cells. In addition, the ERK pathway inhibitor, U0126 (10 μM), significantly reversed the positive effect of TNF-α on the protein levels of RUNX2 and BSP.ConclusionsThe EphB4, TNFR2 and ERK/MAPK signaling pathway comprises a signaling axis to mediate the positive effect of TNF-α on osteogenic differentiation.

Highlights

Low concentrations of tumor necrosis factor-alpha (TNF-α) and its receptor Tumor necrosis factor receptor2 (TNFR2) are both reported to promote osteogenic differentiation of osteoblast precursor cells
We found that the expression levels of RUNX2 and BSP were both significantly higher in Tumor necrosis factor alpha (TNF-α)-treated MC3T3-E1 cells than in the control cells (Fig. 1c-f)
EphB4/TNFR2/ ERK/MAPK signaling pathway comprises a signaling axis to mediate the positive effect of TNF-α on osteogenic differentiation It was previously reported that MAPK signaling pathways play an important role in the osteogenic differentiation [26,27,28]

Summary

Introduction

Low concentrations of tumor necrosis factor-alpha (TNF-α) and its receptor TNFR2 are both reported to promote osteogenic differentiation of osteoblast precursor cells. Tumor necrosis factor alpha (TNF-α) is primarily produced by activated macrophages and lymphocytes, and can be detected in endothelial cells and other cell types [1]. At the late stage of wound healing, TNF-α expression is mainly detected in osteoblasts and other mesenchymal cells [10, 11], indicating that TNF-α actively participates in bone fracture regeneration [9]. According to the literature data and our previous studies, a short-term exposure to low concentrations of TNF-α enhances the osteogenic differentiation of MSCs and osteoblasts, which in turn promotes bone regeneration [12,13,14,15]

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