Hypoxia enhances osteoclastogenesis in periodontal ligament cells via expression of VEGF and RANKL

Introduction: Periodontal ligament (PDL) damage caused by dental trauma can lead to local circulatory disorders. The mechanisms through which PDL cells, once exposed to a transient hypoxic environment, contribute to tissue regeneration or resorption of pathological tooth roots after reoxygenation remain unclear. Therefore, we aimed to examine how changes in oxygen (O₂) concentration affect PDL healing. Materials and methods: Human PDL stem cells (hPDL cells) were cultured under normoxic or hypoxic (20% or 1% O₂ concentration) conditions. Vascular endothelial growth factor (VEGF) and receptor activator of nuclear factor kappa-Β ligand (RANKL) expressions were measured using real-time quantitative polymerase chain reaction or Western blotting. Furthermore, a co-culture of hPDL and osteoclast precursor cells was used to demonstrate the effect of changes in O₂ concentration on osteoclast formation. Results: VEGF expression considerably increased over time under hypoxia compared with normoxia. However, during reoxygenation (24 h hypoxia–24 h normoxia), expression markedly decreased under hypoxia. No significant difference in RANKL expression was observed in both conditions after 24 h; however, it remarkably increased under hypoxia compared with normoxia after 48 h. In the osteoclast formation assay, the number of tartrate-resistant acid phosphatase (TRAP)-positive multinucleated cells considerably increased over time under hypoxia compared with normoxia. Notably, when VEGF expression was reduced using small interfering RNA, the number of TRAP-positive multinucleated cells decreased extensively. Conclusion: Under hypoxic conditions, periodontal ligament cells produce VEGF to promote angiogenesis. However, excessive VEGF production, along with RANKL production, induces osteoclast formation. Osteoclast formation can be suppressed using rapid reoxygenation.