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



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.


Human PDL cells were cultured and identified. 20–100 μg/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.


We found that 20–100 μg/mL of PHT did not affect PDL cells proliferation, whereas 50–100 μg/mL of PHT inhibited cell migration. The 50 or 100 μg/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 μg/mL of PHT. Further, 20–50 μg/mL of PHT increased ALP expression, but 100 μg/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.


MAPK pathway involved in PHT-regulated migration and osteogenic differentiation in human PDL cells.

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Periodontal ligament






Phenylmethane sulfonyl fluoride


Sodium dodecyl sulfate polyacrylamide gel electrophoresis


Polyvinylidene fluoride


Triethanolamine buffered saline solution


Horseradish peroxidase


Enhanced chemiluminescent




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This work was supported by National Natural Science Foundation of China [Grant Nos. 11972067,11572030, 11120101001], National Key R&D Program of China [2017YFC0108505]; the Fundamental Research Funds for the Central Universities; the 111 Project [Grant Number B13003].

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Correspondence to Lisha Zheng or Yubo Fan.

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Jing Na, Lisha Zheng, Lijuan Wang, Qiusheng Shi, Nan Liu, Yuwei Guo and Yubo Fan declare that they have no conflict of interest.

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All human subjects research was carried out in accordance with the Declaration of Helsinki and approved by Beihang University Ethics Committee. This study did not involve any animal research.

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Na, J., Zheng, L., Wang, L. et al. Phenytoin Regulates Migration and Osteogenic Differentiation by MAPK Pathway in Human Periodontal Ligament Cells. Cel. Mol. Bioeng. 15, 151–160 (2022).

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  • Human periodontal ligament cells
  • Phenytoin
  • Migration
  • Osteogenic differentiation
  • ALP
  • MAPK