Epithelia in lung, skin, and kidney are often exposed to fluoride, and tissue damage in lung and kidney due to fluoride is well documented. Nevertheless, the biological effects of fluoride on epithelia are poorly investigated. In the present study, we report effects of sodium fluoride (NaF) on the differentiation of a human epithelial cell line, HaCaT. These cells may serve as a keratinocyte model, because they express a wide spectrum of keratins (Ks), and they associate into stratified tissue-like arrangements along with changes in their keratin pattern. NaF was added to the culture medium at concentrations of 0.5 and 5 mM. Cell proliferation remained intact, but cell functions were altered at high dose, and HSP70 was induced. Reverse transcription-polymerase chain reaction and Western blotting revealed that keratin (K) 15 mRNA and protein expression, associated with stratification of epithelia, were inhibited. Also, expression of keratins typical for terminal differentiation, K1 and K10, was decreased and so was the expression of the K1/10 regulating enhancer binding protein c/EBP alpha. Stratification of HaCaT cells was abolished at high fluoride dose, as assessed by electron microscopy. The changes in keratin expression were not reversed by withdrawal of fluoride. Taken together, NaF at high dose blocked terminal differentiation of HaCaT cells, visible by keratin expression and failing stratification. This effect may disturb tissue formation due to altered cell interactions.