Metformin promotes osteogenic differentiation of human periodontal ligament stem cells via KLF2-mediated activation of miR-181a-5p under lipopolysaccharide stimulation

Periodontal ligament stem cells (PDLSCs) constitute a promising source for successful periodontal regeneration. This study aims to explore roles of metformin, krüppel-like factor 2 (KLF2), and miR-181a-5p in mediating osteogenic differentiation of human PDLSCs (hPDLSCs) following lipopolysaccharide (LPS) stimulation. The osteogenic differentiation potential of hPDLSCs isolated from human premolar root samples were examined by alkaline phosphatase (ALP) staining, ALP activity assay, Alizarin red S staining, and Western blotting of osteogenic markers. Metformin pretreatment at dose of 100 μM significantly resulted in increased ALP activity, elevated protein expressions of osteogenic markers, and more generated mineralized matrix in hPDLSCs with LPS stimulation. KLF2 and miR-181a-5p were found to be increased by metformin pretreatment at dose of 100 μM in hPDLSCs with stimulation but not in hPDLSCs without LPS stimulation. The interaction between the KLF2 and the promoter of miR-181a-5p was noted by the dual-luciferase reporter assay. KLF2 knockdown or miR-181a-5p inhibition notably abrogated the improvements of osteogenic differentiation by metformin pretreatment in LPS-stimulated hPDLSCs. The findings of the study indicate metformin protects hPDLSCs against impaired osteogenic differentiation of hPDLSCs after LPS stimulation by KLF2-mediated activation of miR-181a-5p under inflammation conditions.