Effect of fluoride on osteoclast autophagy through the AKT/mTOR/ULK1 signaling pathway

  Background   Fluoride can induce osteoclasts to secrete a variety of enzymes, which can enhance or weaken the function of bone absorption, resulting in osteolysis or osteogenesis. The process of bone resorption after osteoclast activation is closely associated with autophagy.

  Objective   To investigate the effect of fluoride on osteoclast autophagy through the AKT/mTOR/ULK1 signaling pathway.

  Methods   Osteoclasts induced by RAW264.7 cells were divided into control group, NaF group, CQ group, CQ + NaF group, RAP group, and RAP + NaF group, and NaF, CQ, and RAP were administered at a dose of 10 mg/L, 5 mmol/L, and 5 mmol/L, respectively. After 24 hours, laser scanning confocal microscopy was used for the quantification of autophagosomes in osteoclasts; quantitative real-time PCR was used to measure the transcriptional expression levels of Cathepsin K and TRAP genes, and Western blotting was used to measure the levels of phosphorylated proteins p-AKT, p-mTOR, and p-ULK1 in the AKT/mTOR/ULKl signaling pathway, so as to investigate the role of autophagy in fluoride-exposed osteoclasts.

  Results   Laser scanning confocal microscopy showed that 10 mg/L sodium fluoride solution promoted the production of autophagosomes, with a significant increase in the number of autophagosomes, and sodium fluoride and RAP stimulated autophagy in a synergistic way. Quantitative real-time PCR showed that sodium fluoride reduced the expression levels of Cathepsin K and TRAP genes, and sodium fluoride and CQ had a synergistic effect on the expression of the two genes. Sodium fluoride antagonized RAP to stimulate autophagy and inhibit the upregulation of Cathepsin K and TRAP genes. Western blotting showed that sodium fluoride upregulated the protein expression levels of p-AKT and p-mTOR, and increased the expression of p-AKT, p-mTOR, and p-ULK1 together with RAP in a synergistic way.

  Conclusion   Sodium fluoride solution with a concentration of 10 mg/L can promote osteoclast autophagy, inhibit its bone resorption capacity, and result in osteogenesis, which may be associated with the AKT/mTOR/ULK1 signaling pathway.