Human periodontal ligament stem cells and metformin to enhance periodontal regeneration in rats.

BACKGROUND: Effective periodontal repair and regeneration remain a critical clinical need, requiring strategies that address the complex regeneration of both bone and cementum within the periodontal complex. The objectives of this study were to: (1) develop a novel strategy for periodontal regeneration via human periodontal ligament stem cells (hPDLSCs) and metformin; (2) investigate the metformin regulatory mechanism via the ERK1/2 signaling pathway that promotes periodontal regeneration; (3) evaluate the effectiveness of the combination of hPDLSCs and metformin in promoting the repair of periodontal defects in rats.

METHODS: hPDLSCs were cultured in medium with graded metformin doses (0-600 μM) to determine the optimal concentration via CCK-8 assay, scratch assay and osteogenic/cementogenic differentiation analyses. ERK1/2 phosphorylation, induced by the optimal metformin concentration, and its inhibition by U0126 (10 μM) were analyzed via Western blot to assess its role in promoting osteogenesis and cementogenesis. In vivo, 30 rats with periodontal defects were divided into five groups: (1) Blank control; (2) Matrigel only; (3) Matrigel + metformin; (4) Matrigel + hPDLSCs; (5) Matrigel + Metformin + hPDLSCs. Micro-CT and histological staining were performed at 6 weeks to evaluate periodontal regeneration. Data were analyzed using one-way and two-way ANOVA (p < 0.05).

RESULTS: 100 μM metformin optimally promoted hPDLSCs proliferation, migration, osteogenesis, and cementogenesis, with ALP activity and mineralization increased by 5-fold and 17-fold, respectively, and cementogenic gene expression upregulated to 6-8 times control levels (p < 0.05). The MEK1/2 inhibitor (U0126) decreased metformin's impact on osteogenesis and cementogenesis by 40-50 % (p < 0.05). Compared to the control group, Matrigel+Met+hPDLSCs group enhanced bone regeneration by 3.6 folds, and increased cemental regeneration by 9.5 folds in rats (p < 0.05).

CONCLUSION: This study focused on the dual role of metformin-hPDLSCs combination in promoting both osteogenesis and cementogenesis, addressing the specific needs of periodontal complex regeneration. Additionally, the ERK1/2 signaling is involved in the metformin-induced osteogenesis and cementogenesis.