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Background. The purpose of this study was the evaluation of the potential and mechanism of phenytoin to promote differentiation of human dental pulp stem cells (hDPSC) into odontoblasts/osteoblasts. Methods. Fourth-generation human hDPSC originating from healthy pulp of third molars was cultured in control as well as phenytoin-containing media (PHT) for 14 days. qPCR was applied to detect the expression of DSPP, DMP1, and ALP genes. Western blot analysis was used to confirm the findings. One-way analysis of variance (ANOVA) was used for statistical analysis ( p &lt; 0.05 ). Information about phenytoin was assessed from PubChem database, while targets of phenytoin were assessed from six databases. Drug targets were extracted based on the differentially expressed genes ( logFC ≥ 1 ,  p &lt; 0.05 ) in the experimental group (50 mg/L PHT, 14 days). GO BP and KEGG pathway enrichment analysis on the obtained drug targets was performed and the target protein functional network diagram was constructed. Results. A concentration below 200 mg/L PHT had no obvious toxicity to hDPSC. The expression of DSPP, DMP1, and ALP genes in the 50 mg/L PHT concentration group increased significantly. The WB experiment showed that the protein content of BMP4, Smad1/5/9, and p-Smad1/5 was significantly increased in 50 mg/L PHT in comparison with the NC group (the group without treatment of PHT) at 14 days. Conclusion. Phenytoin has the ability of promoting the differentiation of hDPSC into odontoblasts and osteoblasts. BMP4/Smad pathway, inducing odontogenic/osteogenic differentiation of hDPSC, appears a main process in this context.

Phenytoin Is Promoting the Differentiation of Dental Pulp Stem Cells into the Direction of Odontogenesis/Osteogenesis by Activating BMP4/Smad Pathway