Grape Seed Proanthocyanins Extract Ameliorates the Neurotoxicity Induced by Fluoride Through mTOR/p70s6k Signaling Pathway in Vivo and in Vitro.

Autophagy dysfunction plays an influential role in fluoride-induced neurotoxicity, but the mechanism has not yet been clarified. The aim of this study was to investigate the effects of grape seed proanthocyanidin extract (GSPE) on sodium fluoride (NaF)-treated offspring SD rats and SH-SY5Y cells, and to elucidate the mechanism of mTOR/p70s6k-mediated autophagy signaling pathway action. The results of in vivo experiments showed that different doses of GSPE treatment could alleviate NaF-induced neurological damage, centering on the improvement of learning and memory ability, recovery of autonomous exploration ability, reduction of anxiety-like behaviors, restoration of neuronal structure in hippocampal tissues, and reduction of the expression of apoptotic protein cleaved-PARP. Meanwhile, GSPE significantly promoted autophagy in hippocampal tissues, as evidenced by the blockage of mTOR/p70s6k phosphorylation and the upregulation of LC3-II and downregulation of p62 expression. To further explore the mechanistic issues, we conducted cellular experiments, which suggested that GSPE pretreatment significantly increased cell viability and strengthened autophagy compared with the NaF group. These results were evidenced by the inhibition of mTOR/p70s6k phosphorylation and the upregulation of LC3-II protein. To further investigate the effect of GSPE on the mTOR/p70s6k pathway, the mTOR inhibitor rapamycin (RAPA) and activator MHY1485 were incorporated. The results indicated that GSPE inhibited the enhancement of mTOR phosphorylation in combination with MHY1485, and further inhibited the phosphorylation of mTOR in combination with RAPA, and the levels of LC3-II and p62 and immunofluorescence results also confirmed that the pretreatment of GSPE promoted cellular autophagy. Similarly, Hoechst33342 fluorescence intensity and apoptotic protein cleaved-PARP expression were further reduced after GSPE combined with RAPA or MHY1485. In addition, downstream of the mTOR/p70s6k pathway, brain-derived neurotrophic factor (BDNF), which has been widely used in studies of mood disorders, was significantly reduced by GSPE pretreatment, suggesting that GSPE may not only have a role in attenuating neurotoxicity, but also potentially in improving mood. In conclusion, the present study reveals that GSPE may alleviate fluoride-induced neurotoxicity by inhibiting the activation of the mTOR/p70s6k signaling pathway and thereby enhancing cellular autophagy and antagonizing apoptosis.