The Role of Thymosin Beta 4 on Odontogenic Differentiation in Human Dental Pulp Cells

Sang-Im Lee,Eun-Cheol Kim,Hwa-Jeong Lee,Hee-Jae Cha,Duck-Su Kim,Irina Kerkis

doi:10.1371/journal.pone.0061960

Sang-Im Lee, Eun-Cheol Kim + Show 4 more

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https://doi.org/10.1371/journal.pone.0061960

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Journal: PloS one	Publication Date: Apr 17, 2013
Citations: 36	License type: CC BY 4.0

Affiliation: Kyung Hee University, Kosin University

Abstract

We recently reported that overexpression of thymosin beta-4 (Tβ4) in transgenic mice promotes abnormal hair growth and tooth development, but the role of Tβ4 in dental pulp regeneration was not completely understood. The aim of this study was to investigate the role of Tβ4 on odontoblastic differentiation and the underlying mechanism regulating pulp regeneration in human dental pulp cells (HDPCs). Our results demonstrate that mRNA and protein expression of Tβ4 is upregulated during odontogenic differentiation in HDPCs. Transfection with Tβ4 siRNA decreases OM-induced odontoblastic differentiation by decreasing alkaline phosphatase (ALP) activity, mRNA expression of differentiation markers, and calcium nodule formation. In contrast, Tβ4 activation with a Tβ4 peptide promotes these processes by enhancing the phosphorylation of p38, JNK, and ERK mitogen-activated protein kinases (MAPKs), bone morphogenetic protein (BMP) 2, BMP4, phosphorylation of Smad1/5/8 and Smad2/3, and expression of transcriptional factors such as Runx2 and Osterix, which were blocked by the BMP inhibitor noggin. The expression of integrin receptors α1, α2, α3, and β1 and downstream signaling molecules including phosphorylated focal adhesion kinase (p-FAK), p-paxillin, and integrin-linked kinase (ILK) were increased by Tβ4 peptide in HDPCs. ILK siRNA blocked Tβ4-induced odontoblastic differentiation and activation of the BMP and MAPK transcription factor pathways in HDPCs. In conclusion, this study demonstrates for the first time that Tβ4 plays a key role in odontoblastic differentiation of HDPCs and activation of Tβ4 could provide a novel mechanism for regenerative endodontics.

Highlights

Dental pulp has the capacity to generate reparative dentin in response to damage from infection, exposure, trauma, and chemicals [1]
To investigate the expression of Thymosin beta-4 (Tb4) mRNA and protein during odontoblastic differentiation of human dental pulp cells (DPCs) (HDPCs), cells were cultured in osteogenic medium (OM) for 14 days and the samples analyzed by conventional reverse transcriptase-polymerase chain reaction (RT-PCR) and Western blotting (Figure 1A)
HDPCs showed higher alkaline phosphatase (ALP) activity, mRNA expression of the odontogenic differentiation genes, and mineralized nodule formation when cultured in OM compared with those in control medium, which results confirmed that HDPCs can differentiate into odontoblast-like cells (Fig. 1D–F)

Summary

Introduction

Dental pulp has the capacity to generate reparative dentin in response to damage from infection, exposure, trauma, and chemicals [1]. Reparative dentinogenesis requires the growth and differentiation of dental pulp cells (DPCs) into odontoblasts [2]. The regulation of odontogenic differentiation in human DPCs (HDPCs) has important implications for the development of new therapeutic strategies for vital pulp therapy, but the molecular mechanisms inducing odontoblastic growth and differentiation remain to be elucidated. Tb4 is upregulated during endothelial cell differentiation and has been shown to stimulate angiogenesis by differentiation and directional migration of endothelial cells and tube formation [10]. Tb4 has been shown in various rodent models to promote stem cell migration and differentiation into keratinocytes and hair follicles in the bulge region, inducing dermal repair and causing increased hair growth [8,11]

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