Figure 10. Pyrophosphate regulates cementoblast mineralization and nucleotide pyrophosphohydrolase (NTPPPH) activity, in vitro.

(A) By von Kossa staining, OCCM.30 cells cultured with 5 mM BGP produced mineral nodules by days 6 and 8, while cells receiving only AA did not mineralize. The low dose of 10 µM PP_i did not affect mineral nodule precipitation, however, the higher dose of 100 µM was a potent inhibitor of mineral nodules. (B) Quantitative calcium assay performed on days 6 and 8 confirmed visual mineral nodule staining by von Kossa. (C) Relative ALP enzyme activity was not affected by inhibition of mineralization by 100 µM PP_i. (D) NTPPPHase activity was increased under mineralizing conditions, but inclusion of 100 µM PP_i brought activity back to basal levels of non-mineralizing cultures. Graphs show mean +/− SD for n = 3 samples. Lowercase letters indicate treatment comparison at each time point, where different letters indicate a statistically significant intergroup difference. Uppercase letters indicate comparisons over time in the same treatment group, where different letters indicate a statistically significant intragroup difference. Values sharing the same uppercase or lowercase letter in were not significantly different. Means were compared by ANOVA (p<0.05) followed by the Tukey test for direct pair-wise comparisons.