Platelet-rich Fibrin Promotes the Proliferation and Osteo-/odontoblastic Differentiation of Human Dental Pulp Stem Cells

Abstract

Background: Pulp regeneration is a promising strategy that promotes the continued development of young permanent teeth with immature apical foramen. Platelet-rich fibrin (PRF) was found to stimulate the proliferation and differentiation of osteoblasts, but its effects on osteoblast/odontoblast differentiation of human dental pulp stem cells (hDPSCs) is unknown. Methods: The hDPSCs were isolated and identified by flow cytometry using surface known markers. The CCK-8 assay and the expression of Ki67 and PCNA were used to examine hDPSC proliferation. After 7 days culture in osteo-/odontoblastic induction medium with various concentrations of liquid PRF (0, 10% and 20%), the marker of the early stage of osteogenesis-intracellular alkaline phosphatase (ALP) was checked. After 21 days culture, matrix mineralization was checked using Alizarin Red S and quantified. The mRNA and protein levels of osteo-/odontoblastic genes including RUNX2, DSPP, DMP1 and BSP were measured by qRT-PCR. Notch signal was checked by Western blot to analyze three key proteins (Notch 1, Jagged 1 and Hes 1). Results: PRF-treated groups showed higher expression of Ki-67 and PCNA, higher ALP activity, and the higher dose showed a stronger induction. PRF promoted osteo-/odontoblastic differentiation of hDPSCs indicated by elevated protein levels and mRNA levels of the expression of osteo-/odontoblastic markers. The three key proteins in Notch signaling all showed an increase compared with the control group, and increased as the PRF concentration became higher. Conclusion: PRF can promote the proliferation and osteo-/odontoblastic differentiation of hDPSC, which may be through the Notch signaling pathway.