Previously, we have shown that stem cells from apical papilla (SCAPs) can be chemoattracted by stromal cell–derived factor-1α (SDF-1α). The purpose of this study was to investigate the intracellular signaling pathways involved in SDF-1α–mediated migration of SCAPs.Methods:
Chemotaxis assays were performed to assess the effect of phosphatidylinositol 3-kinase (PI3K) and protein kinase C (PKC) signaling pathways in the SDF-1α–mediated migration of SCAPs using inhibitors of PI3K (LY294002) or PKC (GF109203X). The Cell Counting Kit-8 assay (Dojindo Laboratories, Kumamoto, Japan) was used to evaluate the effect of the inhibitors on the proliferation of SCAPs. The expression of focal adhesion–related proteins was examined by immunofluorescence staining and Western blot analysis. Phosphorylation of PI3K subunit p85 and PKC after SDF-1α induction was evaluated by Western blot.Results:
The inhibition of PI3K or PKC signaling pathways significantly reduced SDF-1α–mediated migration of SCAPs. The inhibitors had no effect on the proliferation of SCAPs. Immunofluorescence analysis revealed that SDF-1α stimulated focal adhesion formation and stress fiber assembly in SCAPs, in addition to up-regulation of the expression of focal adhesion molecules, including p-focal adhesion kinase, p-paxillin, and vinculin. Pretreatment with PI3K or PKC inhibitors before SDF-1α induction significantly inhibited focal adhesion molecule expression. Moreover, increased phosphorylation of p85 and PKC were observed after SDF-1α stimulation, whereas these phosphorylations were down-regulated by the inhibition of PI3K or PKC signaling pathways.Conclusions:
PI3K and PKC signaling pathways appear to be required for SDF-1α–mediated transmigration of SCAPs. These findings provide insights into the signaling mechanisms that underlie SDF-1α–mediated migration of SCAPs.