Stage-Specific Effects of Fibroblast Growth Factor 2 on the Differentiation of Dental Pulp Cells

Abstract

Dentinogenesis is a complex and multistep process, which is regulated by various growth factors, including members of the fibroblast growth factor (FGF) family. Both positive and negative effects of FGFs on dentinogenesis have been reported, but the underlying mechanisms of these conflicting results are still unclear. To gain a better insight into the role of FGF2 in dentinogenesis, we used dental pulp cells from various transgenic mice, in which fluorescent protein expression identifies cells at different stages of odontoblast differentiation. Our results showed that the continuous exposure of pulp cells to FGF2 inhibited mineralization and revealed both the stimulatory and inhibitory effects of FGF2 on the expression of markers of dentinogenesis and various transgenes. During the proliferation phase of in vitro growth, FGF2 increased the expression of markers of dentinogenesis and the percentages of dentin matrix protein 1/green fluorescent protein (DMP1-GFP)-positive functional odontoblasts and dentin sialophosphoprotein (DSPP)-Cerulean-positive odontoblasts. Additional exposure to FGF2 during the differentiation/mineralization phase of in vitro growth decreased the extent of mineralization and the expression of markers of dentinogenesis and of the DMP1-GFP and DSPP-Cerulean transgenes. Recovery experiments showed that the inhibitory effects of FGF2 on dentinogenesis were related to the blocking of the differentiation of cells into mature odontoblasts. These observations together showed the stage-specific effects of FGF2 on dentinogenesis by dental pulp cells, and they provide critical information for the development of improved treatments for vital pulp therapy and dentin regeneration.

Enhanced Dentinogenesis of Pulp Progenitors by Early Exposure to FGF2

“…Cultures were processed for immunocytochemistry for detection of DSPP-Cerulean and phospho-Erk1/2 using anti-GFP (Invitrogen, Grand Island, NY, USA) and rabbit anti-mouse phospho-Erk1/2 (Cell Signaling, Boston, MA, USA) antibodies, respectively, as previously described (Sagomonyants and Mina 2015).…”

“…At different time points, the mean fluorescence intensity in culture wells was measured as previously described (Kuhn et al 2010;Sagomonyants and Mina 2015).…”

“…We have used a series of green fluorescent protein (GFP) reporter transgenic mice that display stage-specific activation of transgenes during odontoblast differentiation in vivo and in vitro to gain a better understanding of the progression of progenitor cells in the odontoblast lineage (Balic et al 2010;Balic and Mina 2011;Sagomonyants and Mina 2015). These studies showed that 2.3-GFP and 3.6-GFP transgenes identify cells at early stages of odontoblast differentiation (polarizing odontoblasts that lack expression of Dmp1 and Dspp; Balic et al 2010), whereas DMP1-GFP and DSPP-Cerulean transgenes identify postmitotic functional and fully differentiated odontoblasts within mineralized nodules, respectively (Balic and Mina, 2011;Mina 2014, 2015).…”

“…These studies showed that 2.3-GFP and 3.6-GFP transgenes identify cells at early stages of odontoblast differentiation (polarizing odontoblasts that lack expression of Dmp1 and Dspp; Balic et al 2010), whereas DMP1-GFP and DSPP-Cerulean transgenes identify postmitotic functional and fully differentiated odontoblasts within mineralized nodules, respectively (Balic and Mina, 2011;Mina 2014, 2015). Using these transgenic animals, we have recently reported that the effects of FGF2 on differentiation of progenitor cells into odontoblasts are stage specific and dependent on the stage of cell differentiation/maturity (Sagomonyants and Mina 2015). Continuous exposure of pulp cells to FGF2 inhibited mineralization and revealed both stimulatory and inhibitory effects of FGF2 on expression of markers of odontoblast differentiation and various transgenes (Sagomonyants and Mina 2015).…”

“…Using these transgenic animals, we have recently reported that the effects of FGF2 on differentiation of progenitor cells into odontoblasts are stage specific and dependent on the stage of cell differentiation/maturity (Sagomonyants and Mina 2015). Continuous exposure of pulp cells to FGF2 inhibited mineralization and revealed both stimulatory and inhibitory effects of FGF2 on expression of markers of odontoblast differentiation and various transgenes (Sagomonyants and Mina 2015). During the proliferation phase of in vitro growth, FGF2 transiently increased expression of markers of odontoblast differentiation, including Dspp and Dmp1.…”