Effect of periostin silencing on Runx2, RANKL and OPG expression in osteoblasts

Normal tooth eruption is closely related to relevant genes and the dynamic balance between osteoblasts and osteoclasts. If secretion of RANKL and OPG by osteoblasts is disordered, relevant gene deficiencies or mutations will result in serious tooth eruption disturbances, e.g., cleidocranial dysplasia (CCD). Thus, we examined changes in Runx2, RANKL, and OPG protein expression in MC3T3-E1 cells after silencing the periostin gene, thus, providing an experimental basis to study tooth eruption mechanisms.

Based on previous research, cells were divided into two groups according to the virus number: the contrast group (NC group; pFU-GW-016 PSC53349-1) and the experimental group (KD group; LVpFU-GW-016PSC66473-1). Cells were infected with the lentiviral vector (multiplicity of infection = 100) and assessed by image cytometry 72 h after infection. After screening cells for the strongest gene silencing effect, Runx2, RANKL and OPG protein expression were detected by western blotting.

Based on quantitative PCR, the periostin gene silencing efficiency in the KD group was over 90% (P < 0.01). After periostin gene silencing, compared with the control group, Runx2 and RANKL expression in the KD group was reduced (P < 0.01 and P < 0.05, respectively), but OPG protein expression showed no significant change (P > 0.05). The RANKL/OPG ratios in the KD group were lower than those in the NC group after periostin gene silencing (P < 0.05).

Silencing periostin may reduce the expression of Runx2, suggesting that there may be a synergistic relationship between periostin and Runx2 in their effects on osteoblast differentiation, while reducing RANKL expression obviously confirms that the NF-κB (nuclear factor κB) pathway plays an important role in this process and that periostin silencing changes the underlying tendency toward bone metabolism. This method could even provide an experimental basis for using exogenous periostin protein to treat some abnormal bone metabolism diseases, as it could be used as a supplement for the treatment of tooth eruption abnormalities caused by Runx2 gene deficiencies or mutations (CCD).