Title: Effects of High Fluoride Exposure on Calcium Homeostasis in LS8 cells
Johnny Groeling (Presenter)
New York University College of Dentistry
Francisco Aulestia, New York University College of Dentistry
Miriam Eckstein, New York University
Rodrigo Lacruz, New York University
Objectives: During enamel development, high fluoride exposure leads to dental fluorosis (DF). The impact of DF varies across geographical zones but in the US it is estimated that 2% of the population present with DF. Previously, it was reported that treatment of the enamel cell line LS8 cells with high fluoride concentration resulted in unfolded protein response (UPR). UPR is an endoplasmic reticulum (ER) stress mediated response that can prevent cell death by handling misfolded proteins and can be activated in conditions of low ER calcium load. Here, we focus on analyzing the effects of excessive fluoride on ER calcium homeostasis and ER associated stress.
Methods: LS8 cells were treated with NaF for 24 hrs. A subset of these cells were loaded with the calcium dye Fura2-AM and stimulated with thapsigargin to passively deplete the ER calcium stores via inhibition of the sarco-endoplasmic reticulum Ca-ATPase (SERCA) pumps. Cells were imaged in the Flexstation III plate reader to determine differences in ER calcium release. Some cells were used to assess changes in UPR via gene expression by real time PCR.
Results: We observed a significant decrease in endoplasmic reticulum calcium load between control and NaF treated cells using thapsigargin. The expression of a number of genes involved in UPR, including GRP78, were upregulated in NaF treated cells, as had also been reported by other groups.
Conclusions: These data indicate that ameloblasts treated with NaF experience increased levels of ER stress and decreased ability to store calcium.