Biochemical and Biophysical Research Communications
Adenosine receptors enhance the ATP-induced odontoblastic differentiation of human dental pulp cells
Introduction
Human dental pulp cells (HDPCs) have attracted the interest of researchers in the field of tissue regeneration because of their accessibility and abundance of stem/progenitor cells [1,2]. Recent studies have reported that HDPCs can differentiate not only into odontoblasts for dentin regeneration, but also into osteoblasts, which can repair bone defects under appropriate conditions [3,4]. The differentiation of mesenchymal cells in HDPCs into odontoblasts is induced by multiple cytokines such as four and a half LIM domains 2 (FHL2) [5], bone morphogenetic protein (BMP2) [6], and ID1 (a downstream target of BMP2 signaling) [7].
Purinergic signaling can regulate the proliferation [8], differentiation [9], and death [8] of different types of stem cells. As critical signaling molecules in this pathway, adenosine triphosphate (ATP) and its hydrolysates act through purinergic receptors, which are classified into P1 (adenosine receptors [ARs], e.g. A1R, A2AR, A2BR, and A3R) and P2 (P2XR, e.g. P2X1–7R; P2YR, e.g. P2Y1,2,4,6,11–14R) receptors. P1 receptors are primarily activated by adenosine, whereas those in the P2 category are mainly regulated by purines such as ATP and adenosine diphosphate (ADP) [10]. Cutarelli et al. [11] were the first to report the biphasic effects of ATP on differentiation and mineralization in human osteoblasts, showing an increase in these processes in response to low concentrations (<100 μM), whereas high concentrations (>100 μM) led to a decrease in these processes, which they suggested was due to the combined activation of P2 receptors and ATP hydrolysis products (e.g. ADP, AMP, adenosine, the mineralization inhibitor PPi, and the mineralization promoter inorganic phosphate [Pi]). In contrast, the osteogenic effects of ATP on human bone marrow mesenchymal stem cells was due to adenosine stimulation of the AR subtype, A2BR [12].
Extracellular ATP and downstream purinergic signaling have also been proposed to contribute to dental pulp tissue healing and dentin regeneration. Mechanical and thermal stimulation of external dentin can induce ATP release in dental pulp through pannexins [13]. Cold stimulation was also reported to induce ATP release from human odontoblast-like cells [14]. Our previous study demonstrated that high concentrations of ATP (800 μM) can induce odontoblastic differentiation of HDPCs, whereas low concentrations (<400 μM) promoted cell proliferation, and that P2 receptors and the ERK/MAPK signaling pathway were involved in this ATP-induced odontoblastic differentiation [15]. These investigations indicated positive roles for ATP and P2 receptors in dental pulp wound healing and dentin formation. However, the specific role of ARs in ATP-induced odontoblastic differentiation of HDPCs, and the effects of adenosine, the hydrolysate of ATP, on HDPC odontoblastic differentiation remain unknown.
All four AR subtypes were demonstrated to be expressed in dental pulp stem cells (DPSCs), and stimulation of A1R might enhance the osteogenic differentiation of DPSCs, moreover, the progress of dentinogenesis is similar to that of osteogenesis, to some extent [16]. We conducted this study to identify the role of ARs in ATP-induced odontoblastic differentiation of HDPCs, and to determine whether the ARs activation by adenosine can induce HDPC odontoblastic differentiation independently, without the induction of ATP.
Section snippets
Cell culture
This study was approved by the Ethics Committee of Peking University School and Hospital of Stomatology (IRB-2013055), and informed consent was obtained. HDPCs were isolated according to a previous report [15]. Cells were cultured in proliferation medium (PM) containing α-minimum essential medium (α-MEM) (Gibco, Grand Island, NY) with 10% fetal bovine serum (Corning Cellgro, NY), 1% L-glutamine (Sigma-Aldrich, St. Louis, MO), and 1% penicillin/streptomycin (Gibco, BRL). For ATP-induced
Expression of adenosine receptor subtypes in HDPCs
To study the expression of ARs in HDPCs, RT-PCR and real-time PCR were performed. The results showed that all four AR subtypes, A1R, A2AR, A2BR, and A3R, were expressed in HDPCs (Fig. 1A). However, the levels of A2AR and A2BR were much higher than those of A1R and A3R. A2BR exhibited the highest and A3R the lowest expression levels (Fig. 1B). These results were consistent with those of a previous study [16].
Adenosine receptor subtypes were activated in HDPCs exposed to ATP
To investigate the response of ARs to ATP exposure, the mRNA and protein expression
Discussion
Purinergic signaling affects various physiological processes, including osteogenesis [11], neurogenesis [9], inflammation [21], and pain [22]. Receptors involved in this signaling are classified into two main groups: P1 (adenosine receptors [ARs]) and P2 receptors. In our previous study, we had demonstrated that ATP could induce odontoblastic differentiation of HDPCs by analyzing the key events in this process, including DMP1 and DSPP expression, as well as the mineralization capacity of HDPCs
Conflict of interest
The authors declare no conflicts of interest.
Acknowledgments
This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors. We would like to thank Editage [www.editage.cn] for English language editing.
References (27)
- et al.
Prospects for translational regenerative medicine
Biotechnol. Adv.
(2012) - et al.
Expression of Pannexin3 in human odontoblast-like cells and its hemichannel function in mediating ATP release
Arch. Oral Biol.
(2015) - et al.
Effects of adenosine triphosphate on proliferation and odontoblastic differentiation of human dental pulp cells
J. Endod.
(2016) - et al.
Adenosine A1 receptor stimulation enhances osteogenic differentiation of human dental pulp-derived mesenchymal stem cells via WNT signaling
Stem Cell Res.
(2013) - et al.
Effects of WNT10A on proliferation and differentiation of human dental pulp cells
J. Endod.
(2014) - et al.
The G(s)-coupled adenosine A(2B) receptor recruits divergent pathways to regulate ERK1/2 and p38
Exp. Cell Res.
(2003) - et al.
A2B adenosine receptor promotes mesenchymal stem cell differentiation to osteoblasts and bone formation in vivo
J. Biol. Chem.
(2012) - et al.
Dental pulp of the third molar: a new source of pluripotent-like stem cells
J. Cell Sci.
(2012) - et al.
Stem/progenitor cell-mediated de novo regeneration of dental pulp with newly deposited continuous layer of dentin in an in vivo model
Tissue. Eng. Part A
(2010) - et al.
Promising cell-based therapy for bone regeneration using stem cells from deciduous teeth, dental pulp, and bone marrow
Cell Transplant.
(2011)
FHL2 mediates tooth development and human dental pulp cell differentiation into odontoblasts, partially by interacting with Runx2
J. Mol. Histol.
Bone morphogenetic protein 2-induced human dental pulp cell differentiation involves p38 mitogen-activated protein kinase-activated canonical WNT pathway
Int. J. Oral Sci.
Id1 expression level determines the differentiation of human dental pulp stem cell
J. Dent. Res.
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