Epigenetic Regulation of Osteoblastic Differentiation of Periodontal Ligament derived-cells

Objective: The human periodontal ligament derived-cells (HPDL cells, Lonza) have a potential ability to form mineralization in vitro. The osteoblastic differentiation requires the coordinated stepwise expression of multiple genes. Epigenetic modifications are often involved in the transcriptional level of mRNAs. The present study investigated the role of epigenetic regulation in the osteoblastic differentiation of HPDL cells in response to lipopolysaccharide (LPS) derived from Porphyromonas gingivalis (P. g). Methods: HPDL cells were grown in DMEM supplemented with 10% fetal bovine serum. The cells were incubated with LPS (P. g, InvivoGen) at the concentration of 0.1, 1 and 10 µg/ml for 24 hrs. In a part of the experimental series, the cells are treated with both DNA methyltransferase inhibitors, 5-aza-dCyd (5-aza, Sigma) or histone deacetylase inhibitor, Trichostatin A (TSA, Sigma) and LPS. No treatments were used as controls. Expression levels of histone deacetylase (HDAC1), DNA methyltransferase (DNMT1), Type I collage, bone sialoprotein (BSP), osteopontin (OPN) and osteocalcin (OCN) mRNAs were evaluated by RT-PCR and quantitative Real-Time PCR using TaqMan probes. The significance of the differences was analyzed using Student's t-test (n=5). Results: Each expression level of HDAC1, DNMT1, BSP, OPN and OPC was significantly lower in the cell treated with LPS than in control. When the cells were treated both LPS and 5-aza or TSA, there was no significant difference between the experimental and control groups. Conclusion: The results indicate that epigenetic modification may affect osteoblastic differentiation in periodontal ligament derived-cells.