Figure 2.

Policosanol increases osteoblast differentiation through AMPK activation. MC3T3-E1 cells were treated with 50 μg/mL policosanol and/or 1 μM Com. C. (a) Cells were harvested to determine the protein levels. Western blotting assay was performed using the indicated antibodies. (b) Policosanol- and Com-C-treated cells were cultured for 4 days. qPCR was performed to analyze the expression of osteogenic genes. (c) Western blotting assay was performed using the indicated antibodies. (d) ALP staining was performed to estimate ALP activity as an indicator of osteoblast differentiation after treatment with policosanol and Com C. (e) MC3T3-E1 cells were treated with policosanol and Com C for 21 days. The cells were stained with ARS solution to reveal extracellular mineralization. * p < 0.05, ** p < 0.01 compared with the untreated control; ^# p < 0.05, ^## p < 0.01 compared with the policosanol-treated group. Data represent the mean ± SD of three individual experiments.