Hypoxia Regulates the Proliferation and Osteogenic Differentiation of Human Periodontal Ligament Cells Under Cyclic Tensile Stress via Mitogen‐Activated Protein Kinase Pathways

Previous studies have shown that periodontal ligament exists in a hypoxic microenvironment, especially under the condition of periodontitis or physical stress. The present study is designed to investigate the effects and mechanisms of hypoxia on regulating the proliferation and osteogenic differentiation of human periodontal ligament cells (hPDLCs) under cyclic tensile stress (CTS).hPDLCs were cultured in 2% O2 (hypoxia) or 20% O2 (normoxia) and then subjected to a cyclic in-plane tensile deformation of 10% at 0.5 Hz. The following parameters were measured: 1) cell proliferation by flow cytometry; 2) cell ultrastructure by transmission electron microscopy; 3) expression of hypoxia-inducible factor-1α (HIF-1α) and osteogenic relative factors (i.e., secreted phosphoprotein 1 [SPP1; also known as bone sialoprotein I/osteopontin], runt-related transcription factor 2 [RUNX2], and transcription factor Sp7 [SP7]) by real-time polymerase chain reaction and Western blot; and 4) involvement of mitogen-activated protein kinase (MAPK) signaling pathways by Western blot with specific inhibitor.Proliferation index in the hypoxia with CTS group was significantly higher than in other groups. Significant increases in HIF-1α, SPP1, RUNX2, and SP7 occurred in the presence of hypoxia for 24 hours. In addition, MAPK inhibitor (PD 98,059) significantly attenuated hypoxia and CTS-induced phosphor-ERK1/2 (extracellular regulated kinase 1/2), phosphor-JNK (c-jun N-terminal kinase), and phosphor-P38 expression.Hypoxia regulates CTS-responsive changes in proliferation and osteogenic differentiation of hPDLCs via MAPK pathways. Hypoxia-treated hPDLCs may serve as an in vitro model to explore the molecular mechanisms of periodontitis.