Figure 4

FO activates osteoblast-specific protein expression and mineralization/calcification in osteosarcoma MG-63 human osteoblast-like cells. (A) MG-63 cells (3 × 10³ cells/mL) were seeded overnight and then treated with FO (50 µg/mL and 100 µg/mL) or DEX (100 nM) for 7 days. Protein was extracted and western blotting analysis was performed using each specific antibody. β-Actin was used as the internal control of protein expression. All protein expression was normalized by the density of β-actin. The nuclear localization of runt-related transcription factor 2 RUNX2 (B) and osterix OSX (C) was measured by immunostaining. (D) ALP activity (mineralization) was measured using a TRACP & ALP assay kit. (E) In vitro calcification was detected by alizarin red staining. Significant differences among the groups were determined using the one-way ANOVA followed by Bonferroni correction. All data are presented as mean ± SEM (*** p < 0.001, ** p < 0.01, and * p < 0.05 versus untreated group). FO; fermented extract of C. gigas and DEX; dexamethasone. FO; fermented extract of C. gigas and DEX; dexamethasone.