The objective of the present work was to investigate the effect of Periostin (POSTN) silencing on autophagy in osteoblasts, and provide an experimental basis for studying the mechanism of dental eruption. The cells were divided into the following four groups according to their viral number: the NC group, pFU-GW-016PSC53349-1; group KD1, LVpFU-GW-016PSC66471-1; group KD2, LVpFU-GW-016PSC66472-1; and group KD3, LVpFU-GW-016PSC66473-1. The lentiviral vector was infected at MOI = 100 in the ENi.S medium containing 5 g/mL Polybrene. The target gene expression was observed by a Celigo® Image Cytometer at 72 hours after infection, and the positive rate of fluorescence was noted. A two-step method of quantitative real-time PCR (qRT-PCR) was used to detect the silencing effect of POSTN. Western blotting was then performed to assess the expression of autophagy-related proteins Beclin-1 and LC3 in the group showing the best gene silencing effects. The experimental results showed that there was strong green fluorescence in group KD3. As confirmed via qRT-PCR analysis, the POSTN silencing efficiency in group KD3 reached 92.1%. The Western blotting revealed that the expression of Beclin-1 protein in group KD3 was significantly higher than that in the NC group. However, the LC3 protein expression was not significantly different from that of the control group. The lentiviral vector targeting POSTN in osteoblasts was constructed successfully. In addition, the expression of autophagy protein in mouse osteoblasts increased after POSTN silencing. This finding may provide new approaches for understanding the molecular signal transduction of POSTN during the tooth eruption process.
Keywords: silencing; Beclin1; LC3; osteoblasts.