Fluoride at mitogenic concentrations increases the steady state phosphotyrosyl phosphorylation level of cellular...

This study was designed to test the hypothesis that treatment of human bone cells with mitogenic concentrations of fluoride would lead to an increase in the steady state level of tyrosyl phosphorylation of specific cellular proteins. With an immunoblot assay method, it was found that mitogenic concentrations of fluoride (i.e. 50-200 mumol/L) induced a dose- and time-dependent increase in the level of tyrosyl phosphorylation of at least 13 cellular proteins in both normal human bone cells and human TE85 osteosarcoma cells. Time-course studies revealed that a statistically significant increase in tyrosyl phosphorylation of these 13 cellular proteins in human bone cells was observed after 3-6 h of fluoride treatment and was sustained for up to 24 h. This time course was not compatible with a direct activation of tyrosyl kinases, as epidermal growth factor, which activates tyrosyl kinase activity, induced an immediate and acute response that was rapidly reversible within 1 h. Although fluoride increased the steady state tyrosyl phosphorylation of the cellular proteins in human bone cells, the same micromolar doses of fluoride had no effect on human skin fibroblasts, which are fluoride-nonresponsive cells. The effects of fluoride were rapidly reversible in the absence of fluoride and could be acutely potentiated by pretreatment with epidermal growth factor. In summary, we have shown for the first time that mitogenic concentrations (i.e. 50-200 mumol/L) of fluoride increased the steady state level of tyrosyl phosphorylation of at least 13 cellular proteins in human bone cells, and that the increases were relatively show in onset and sustained. In conclusion, these findings are consistent with the hypothesis that the osteogenic actions of fluoride are mediated at least in part by an inhibition of the activity of one or more fluoride-sensitive phosphotyrosyl protein phosphatases in human bone cells.