Lipopolysaccharide inhibits osteogenic differentiation of periodontal ligament stem cells partially through toll-like receptor 4-mediated ephrinB2 downregulation.

Abstract

This study aimed to investigate the possible crosstalk between LPS/toll-like receptor 4 (TLR4) and ephrinB2 signaling in mediating osteogenic differentiation of PDLSCs. Human periodontal ligament stem cells (hPDLSCs) were harvested and treated with different concentrations of LPS under osteogenic induction. qPCR, alkaline phosphatase (ALP) staining, and Alizarin Red S staining were performed to assess osteogenic gene expression, ALP activity, and mineralized nodule formation. EphrinB2 mRNA and protein expressions after LPS treatment were also determined. To explore the role of ephrinB2 in LPS-impaired osteogenic differentiation of hPDLSCs, hPDLSCs were stimulated with ephrinB2-Fc or transfected with ephrinB2 lentivirus, and then, the osteogenic differentiation capacity was evaluated. LPS inhibited osteogenic differentiation of hPDLSCs and downregulated ephrinB2 expression in hPDLSCs during osteogenic differentiation. Blockage of TLR4 partially reversed LPS-induced decrease in ephrinB2 expression. EphrinB2-Fc promoted mineralized nodule formation and increased the expression of ALP, osteocalcin (OCN), and bone morphogenetic protein 2 (BMP2) in hPDLSCs. EphrinB2-overexpressing hPDLSCs treated with LPS expressed higher ALP and BMP2 mRNA and higher ALP activity and showed more mineralized nodule formation, when compared with wide-type hPDLSCs treated with LPS. Our data suggested that LPS decreased the osteogenic differentiation capacity of hPDLSCs partially through downregulation of ephrinB2 expression via LPS/TLR4 signaling. Upregulation of ephrinB2 partially reversed the impaired osteogenic potential of hPDLSCs induced by LPS. Our results provided a new insight of mechanism underling LPS-mediated osteogenic differentiation inhibition of PDLSCs and clarified a potential target for the management of periodontitis.