FIG. 2.

Calcium block does not rescue OS differentiation in depolarized hMSCs. hMSCs were treated with LaCl₃ to block calcium flux during OS differentiation. (A) IBSP gene expression was quantified by qPCR after 7 days of hMSC culture in OS medium with 40 mM K⁺ (HK) and/or 500 μM LaCl₃ (La). Data points are mean normalized expression ± standard deviation, n = 3. Different letters over the bars represent statistically different groups as determined by one-way ANOVA and the Tukey–Kramer post hoc test (p < 0.05). Groups (a), (b), and (c) were statistically different from one another, but the samples within group (a) were not statistically different from one another. (B) Total calcium content was quantified after 21 days of hMSC culture under the same conditions. Data points represent mean calcium concentration (μg/mL) ± standard deviation, n = 6. Different letters over the bars represent statistically different groups as determined by one-way ANOVA and the Tukey–Kramer post hoc test (p < 0.05). Groups (a), (b), and (c) were statistically different from one another, but the samples within group (a) were not statistically different from one another. (C–F) Representative images of cells stained at day 21 for ALP (blue) and calcified mineral (Alizarin Red S). Cells were grown in media supplemented with La+HK (C), La (D), HK (E), or in normal OS medium (F). Scale bar = 200 μm. HK, high K⁺; IBSP, integrin-binding sialoprotein; LaCl₃, lanthanum chloride; OS, osteogenic.