Figure 5.

SMA 12b downregulates OC-specific gene transcription and induces an antioxidant response during OC differentiation in vitro. Non-adherent BM, from C57BL/6 mice, was harvested and cultured for 4 days with 30 ng/ml M-CSF and 50 ng/ml RANKL (M + R), at which point medium was refreshed and cells were incubated in medium or medium containing 5 µg/ml of SMAs 11a, 12b, or 19o. At day 5 of culture, OCs were lysed, mRNA extracted, and cDNA synthesized. Expression of nfatc1 (A), mmp9 (B), ctsk (C), dc-stamp (D), itgb3 (E), hmox-1 (F), gclc (G), and gclm (H) mRNA was determined by qRT-PCR to assess the phenotypic profile of OCs. Likewise, on day 5 of culture, following the 24-h treatment with SMAs, OCs (I) were stained using the fluorescent stain DCF-DA to measure the presence of reactive oxygen species (ROS). Data are collated from three experiments performed in triplicate, normalized to the M + R control and presented as mean ± SD. One-way ANOVA with Dunn’s post-test was used to analyze experimental conditions compared to M + R, where *p < 0.05, **p < 0.01, and ***p < 0.001.