Phenytoin Regulates Migration and Osteogenic Differentiation by MAPK Pathway in Human Periodontal Ligament Cells

Introduction Periodontal healing requires an adequate number of periodontal ligament (PDL) cells to rebuild the impaired tissue. Phenytoin (PHT) has been reported to promote wound healing and extracellular matrix deposition, which indicates its promising application of periodontal healing. However, the effects of PHT on PDL cells behavior and the underlying mechanism are still unknown. Methods Human PDL cells were cultured and identified. 20-100 mu g/mL PHT were used in our study. The proliferation of PDL cells was determined by the EdU assay. A wound healing assay was used to detect cell migration. Matrix metalloproteinase (MMP)-1, MMP-2, tissue inhibitor of metalloproteinase (TIMP)-1 and TIMP-2 expression were analyzed by real time-PCR. The protein expression of MMP-1 and phosphorylated mitogen-activated protein kinases (MAPKs) were detected by western blotting assay. Osteogenic differentiation was assessed by alkaline phosphatase (ALP) staining. Results We found that 20-100 mu g/mL of PHT did not affect PDL cells proliferation, whereas 50-100 mu g/mL of PHT inhibited cell migration. The 50 or 100 mu g/mL of PHT decreased the gene and protein expression of MMP-1, but increased the gene expression of TIMP-1. MMP-2 and TIMP-2 were not affected by 20-100 mu g/mL of PHT. Further, 20-50 mu g/mL of PHT increased ALP expression, but 100 mu g/mL of PHT depressed ALP expression. The extracellular regulated kinase (ERK), c-Jun N-terminal kinase (JNK), and p38 were activated by PHT. JNK and ERK are involved in PHT-regulated migration. JNK plays an essential role in PHT-induced osteogenic differentiation. Conclusions MAPK pathway involved in PHT-regulated migration and osteogenic differentiation in human PDL cells.