Abstract
The multipotent nature of dental pulp stem cells (DPSCs) promises regenerative endodontic potentials. Alterations in microenvironment have been shown to control the differentiation phenotypes of DPSCs. Understanding the biological mechanisms and finding the optimal DPSC differentiation protocols are crucial for successful DPSC engineering strategies in pulp and dentin healing. The aim of this study is to identify the role of p38 mitogen-activated protein kinase (p38) under normal and oxygen-deprived conditions (2%) to reveal its effect on odontogenic DPSC differentiation. Human DPSCs were isolated from healthy molars and underwent odontogenic differentiation in regular and osteogenic media treated with SB203580, a p38 inhibitor, for 72 hours, and then swapped with osteogenic media for 21 days under hypoxic condition. Immunochemistry and PCR analysis for the various odontogenic differentiation genes and proteins were performed. Our PCR data demonstrate that p38 inhibition resulted in a significant upregulation in odontogenic gene expressions such as DMP-1, DSPP, RUNX, and OSX in normal conditions. Under hypoxia, this effect was reversed. These results were further supported by DSPP immunohistochemistry. The DSPP expression under hypoxia was significantly weaker compared to the control. Our results indicate that p38 represents a negative regulator of the odontogenic DPSC differentiation in normoxia. Under hypoxia, p38 exerts a positive function of DPSC differentiation. Taken together, we identified the p38 and oxygen level as crucial factors to control odontogenic DPSC differentiation providing their essential roles in designing for successful pulp-dentin complex engineering strategies.
Highlights
In recent years in the regeneration field of dentistry, much attention has been given to stem cell biology as they demonstrate the remarkable translational capacity of regenerating dentin and damaged tooth [1, 2]
I Odontogenic Lineage Cell Markers Such as dentin sialophosphoprotein (DSPP), dentin matrix protein (DMP)-1 and RUNX2 are Expressed in the Differentiating and Differentiated Cells from Dental pulp stem cells (DPSCs)
To demonstrate specific in vitro role of p38 in DPSC odontogenic differentiation both in normal and oxygen-deprived conditions, isolated DPSCs were differentiated into odontoblast-like cells using p38 inhibitor SB203580 by our modified differentiation protocol
Summary
In recent years in the regeneration field of dentistry, much attention has been given to stem cell biology as they demonstrate the remarkable translational capacity of regenerating dentin and damaged tooth [1, 2]. Several studies, including ours, have shown that human DPSCs can successfully differentiate into odontoblast-like cells under special culture conditions [3, 4]. These differentiated cells were able to produce dentin. Changes in the cultural micro-environment have been shown to affect the phenotypes of stem cells. Extracellular environments, such as extracellular matrix, secreted factors and direct cell to cell contact, have both positive and negative roles in stem cell differentiation and cell fate determination [5, 6]. The natural oxygen level in the body is remarkably low – approximately 3%
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