FIGURE 5.

Osteoblast-like cells from female Prkca^−/− mice have an enhanced differentiation state in vitro. CLBObs were derived from female WT and Prkca^−/− mice. A, growth curves were determined by counting the cell number at the indicated time points. B, CLBObs derived from WT and Prkca^−/− mice were cultured for the indicated period of time with or without treatment with osteogenesis induction medium (OIM). Alkaline phosphatase activity was determined and normalized to total protein content (n = 12). Unless indicated, comparisons are relative to vehicle-treated cultures of the same genotype at the same time point. C, cultures from WT and Prkca^−/− mice were treated with the PKC activator PMA. Alkaline phosphatase activity was determined normalized to total protein content, and the percentage change in activity relative to vehicle-treated controls is shown. D, quantification of the proportion of culture area stained with alizarin red after 21 days of treatment with osteogenesis induction medium (n = 12). E, representative cultures from WT and Prkca^−/− mice fixed following 21 days of treatment with vehicle (veh) or OIM and stained with alizarin red. F, qRT-PCR quantification of osteoblastic differentiation markers in CLBObs after 14 days of culture (n = 12). β2-MG housekeeping gene expression is shown per μg of RNA. Bars, mean ± S.E. (error bars). *, p < 0.05; **, p < 0.01; ***, p < 0.001 versus WT controls. ###, p < 0.001 versus the percentage change in WT cultures.