Effects of Estradiol on the Proliferation and Odontoblastic Differentiation of Human Dental Pulp Cells 1

Abstrac

Estradiol, an endogenous estrogen could positively influence the proliferation or differentiation of neural stem/progenitor cells. However, the biological effects of estradiol in human dental pulp cells (HDPCs) have not been well studied. The purpose of this study is to investigate the effects of estradiol on the proliferation and odontoblast/osteoblast-like differentiation in HDPCs. For differentiation experiments, cells were cultured in differentiation inductive medium including 100 μM/L ascorbic acid and 10 mM/L β-glycerophosphate. Cell viability, proliferation, levels of mRNA for differentiation-related genes, alkaline phosphate (ALP) activity, and mineralization in the HDPCs on various concentrations of estradiol were assessed. Estradiol stimulated cell viability in a dose-dependent manner. BrdU positive cells and expression of proliferating cell nuclear antigen (PCNA) were increased in estradiol-treated HDPCs, compared with the control. In differentiation inductive medium condition, cell viability of estradiol-treated HDPCs was decreased dose-dependently. Estradiol increased the ALP activity and enhanced formation of mineralized nodule. Moreover, estradiol upregulated ALP, bone sialoprotein (BSP), dentin matrix protein-1 (DMP-1) and dentin sialophosphoprotein (DSPP) in HDPCs compared with untreated control. In addition, ERK was markedly activated in estradiol-treated HDPCs.                 

These findings suggest that estradiol promotes proliferation and odontoblastic differentiation of HDPCs via ERK signaling pathway.

 

Introduction

 

Human dental pulp cells (HDPCs) are isolated from dental pulp tissues, which are located in the centre of a tooth. They are known as stem cells with a high capacity for proliferation and multi-potency, and these cells have the ability to differentiate into odontoblast-like cells and form dentin-like structures (Gronthos et al., 2000). Human dental pulp (HDP) healing and repair are the result of successive and interrelated processes, including the proliferation, chemotaxis, and differentiation of dental pulp cells into odontoblasts leading to reparative dentin formation (Gronthos et al., 2002). Thus, differentiated and undifferentiated cells within dental pulp may contribute to the dentinal regeneration process (Agata et al., 2008). Odontoblasts secrete several collagenous and noncollagenous proteins, including osteonectin, osteopontin (OPN), bone sialoprotein (BSP), dentin matrix protein-1 (DMP-1), and dentin sialophosphoprotein (DSPP) (Butler et al., 1995), which have been used as mineralization markers for the odontoblast-/osteoblast-like differentiation of HDPCs (Gronthos et al., 2000).

There are many factors that influence on proliferarion and differentiation by regulating differentiated or undifferentiated pulp cells. Estrogens (female estrogenic hormones such as  17β-estradiol) are candidates for extrinsic regulators. In a previous study, both 17β-estradiol (E2), known to be an endogenous estrogen, and bisphenol A (BPA), known to be a xenoestrogen, could positively influence the proliferation or differentiation of neural stem/progenitor cells (NS/PCs) (Okada et al., 2010). In addition, in cultured rat PDL cells, estrogen was reported to stimulate the bone forming capacity by increasing alkaline phosphatase (ALP) activity, osteocalcin (OCN) production and the formation of mineralized nodules (Morishita et al.,1999). However, the effect of estrogen on the proliferation and odontoblastic differentiation of HDPCs has not been fully investigated. Mitogen-activated protein kinases (MAPKs) are essential components of the signal transduction system and play critical role in cellular responses including growth, proliferation, differentiation and apoptosis in eukaryotic organisms (Kratchmarova et al., 2005). There are three central elements of MAPK cascades : extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 kinases (Johnson et al., 2002). Although recent studies have found that MAPK signaling is involved in dental pulp cell differentiation and tooth development (Lee et al., 2010; Simson et al., 2009), the intracellular signaling mechanisms responsible for the  regulation of odontoblastic differentiation induced by estrogen in HDPCs are not clear.  

Therefore, the aim of this study is to investigate the effects of estradiol on the proliferation and odontoblastic differentiation of HDPCs and to identify the signal transduction pathways involved in the proliferation and differentiation of HDPCs.