Unraveling the Mechanisms of TMEM16A in Fluoride Toxicity and Tooth Development

Background: Fluorosis is a health condition caused by excessive fluoride intake, which can harm the body, especially the teeth. In this study, we investigated whether transmembrane protein 16A (TMEM16A) plays a role in tooth damage caused by fluorine and whether calcium supplementation can mitigate this damage in mice with dental fluorosis.Methods: Fluorosis in mice was induced by drinking water containing 100 mg/L of sodium fluoride (NaF) and administering different doses of calcium for 10 weeks. The mice were categorized into four groups: the Model group, the Low-Calcium group, the Medium-Calcium group, and the High-Calcium group. We then assessed the effects of calcium supplementation on body weight, fluoride levels in blood, incisors, and bones. We evaluated tooth development-related proteins (bone morphogenetic protein 2 (BMP-2), transformation growth factor beta (TGF-beta), nuclear factor kappa B (NF-kappa B), BCL-2-associated X protein (BAX), and B-cell lymphoma-2 (BCL-2)), and TMEM16A expression using techniques such as quantitative polymerase chain reaction (qPCR), Western blot, hematoxylin and eosin (HE) staining, and immunohistochemistry. We also examined the effect of the TMEM16A inhibitor T16Ainh-A01 on tooth development-related proteins by supplementing it in the calcium administration group. Various doses of NaF (0 mg/L, 50 mg/L, 100 mg/L, 150 mg/L) were given to evaluate the impact of NaF on dental germ development.Results: Compared to the model mice, those receiving medium and high calcium supplementation displayed increased body weight (p < 0.01), reduced fluoride levels (p < 0.01), and decreased apoptotic enamel cells (p < 0.01). TMEM16A, NF-kappa B, and BAX expression levels were higher in the model group but decreased following calcium treatment. Moreover, calcium treatment significantly boosted BMP-2, TGF-beta, and BCL-2 expression levels. When combined with the TMEM16A inhibitor T16Ainh-A01, medium-dose calcium further enhanced BMP-2 and TGF-beta expression while decreasing NF-kappa B expression. As NaF concentration increased, a corresponding rise in fluoride content was observed in blood, tooth germs, and bone tissues. Notably, tooth germ development was delayed and slowed in the NaF-treated group compared to the control group, with TMEM16A expression increasing alongside higher NaF concentration and tooth germ development stage.Conclusions: High concentrations of NaF lead to elevated TMEM16A expression, activating the calcium-activated chloride channel and causing tooth damage. Calcium supplementation or inhibition of TMEM16A expression shows promise in alleviating tooth damage caused by fluorosis, contributing to dental repair in fluoride-affected mice. These insights offer potential therapeutic strategies for managing dental fluorosis.