Abstract
Fluoride (sodium fluoride) is thought to be essential in the development of tooth, and research shows that fluoride can modulate the differentiation of dental stem cells. However, the effects of fluoride on the committed differentiation of stem cells from apical papilla (SCAPs) and the underlying mechanisms remain unclear. Here, SCAPs were isolated from healthy extracted human third molars with immature roots and then were cultured with NaF conditioned media. Cell Counting Kit-8, EdU staining, and flow cytometry were performed to detected the proliferation activity. Alkaline phosphatase (ALP) activity, Alizarin Red staining, Western blot assay, and real-time reverse-transcription polymerase chain reaction were applied to assess the osteo/odontogenic differentiation NaF-treated SCAPs. Western blot assay and transmission electron microscope were used to evaluate the autophagy involved in the differentiation of SCAPs. ALP activity, ALP protein, and messenger RNA (mRNA) expression showed that 0.5 mM was the optimal concentration for the induction of SCAPs by NaF. 0.5 mM NaF-treated SCAPs induced more mineralized nodules as compared with untreated cells. Moreover, the osteo/odontogenic markers (RUNX2, OSX, DSP, and OCN) in mRNA levels were upregulated while the protein levels of these markers increased considerably in 0.5 mM NaF-treated SCAPs. Furthermore, the autophagy-related proteins (LC3, ATG5, and Beclin1) increased in NaF-treated SCAPs, and the osteo/odontogenic makers significantly decreased while silencing ATG5 to block autophagy. In all, sodium fluoride can regulate the osteo/odontogenic differentiation of SCAPs by modulating autophagy.