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

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.

Expression of thymosin beta-4 in human periodontal ligament cells and mouse periodontal tissue and its role in osteoblastic/cementoblastic differentiation

“…In the present study, Tβ4 mRNA and protein expressions were time-dependently upregulated and widely expressed throughout the differentiation process of hPDLCs, which is similar to our recent findings in hDPCs (Lee et al, 2013). However, the expression of Tβ4 mRNA and protein was significantly increased on day four, and decreased from days 14 to day 28 during the differentiation of MDPC-23 cells (Choi et al, 2012).…”

“…To induce differentiation, cells were cultured with osteogenic medium (OM; 50 μg/mL ascorbic acid, 10 mM β-glycerophosphate, and 10 À 7 M dexamethasone) as described previously Kook et al, 2009;Lee et al, 2011;Chung et al, 2014). The cells were then treated with Tβ4 peptide (Abcam) for 14 days as previously described (Lee et al, 2013).…”

“…In contrast, Tβ4 inhibition by Tβ4 siRNA attenuated odontoblastic differentiation in the odontoblast-like cell line, MDPC-23 (Choi et al, 2012). Moreover, we recently demonstrated that odontoblastic differentiation was enhanced by the activation of Tβ4 by Tβ4 peptide, but decreased by Tβ4 siRNA in human dental pulp cells (hDPCs) (Lee et al, 2013). In addition, Tβ4 overexpressing non-odontogenic HaCaT cells showed upregulation of odontogenesis-related gene expressions and had increased calcium deposition (Kiyoshima et al, 2014).…”

Thymosin β4 induces proliferation, invasion, and epithelial-to-mesenchymal transition of oral squamous cell carcinoma

“…Changes in tyrosine phosphorylation of paxillin have been linked to invasion (Summy and Gallick 2003;Badowski et al 2008). Recently, Tβ4 activation has been shown to induce the phosphorylation of paxillin in human dental pulp cells (Lee et al 2013).…”

Expression of Thymosin β4 in Ameloblasts during Mouse Tooth Development

“…In cells of the odontoblastic cell line MDPC-23, the suppression of Tβ4 expression was found to remarkably decrease the expression of DSPP, BSP, OCN, ON, and Col I in the regulation of dentin mineralization and the formation of the mineralized nodule (Choi et al, 2012). Suppression of Tβ4 expression in human dental pulp cells also remarkably reduced the differentiation of odontoblasts and the mineralization, as determined based on the decrease in the expression of alkaline phosphatase, osteopontin, OCN, dentin matrix protein-1, and DSPP (Lee et al, 2013). Tβ4-overexpressing mice show amelogenesis imperfecta due to the reduction in cell polarity caused by the inhibition of actin polymerization that is required for enamel matrix secretion in and migration of presecretory ameloblasts (Cha et al, 2010).…”