Fig. 3

HDAC6 accumulation and histone hypoacetylation on Runx2 promoter in BMSCs from aged mice. BMSCs were isolated from 6- (Young) and 18-month-old (Aged) mice respectively and induced to undergo osteogenesis differentiation. At week 1, total RNA was extracted and subjected to quantitative RT-PCR analysis using primers for Runx2 (A). β-actin was used as an internal control. ChIP was employed to evaluate phosphorylated RNA polymerase II occupancy of the Runx2 promoter in young and aged BMSCs with osteogenic induction (B). Histone samples were collected. Global histone modification levels including acetylated H3K9/K14 and H4K12, and HDAC family members were determined by western blot (C). Histone 3 and 4 were used as loading controls. ChIP assays were used to examine acetylated H3K9/K14 (D), acetylated H4K12 (E), and HDACs (F) occupancy in the Runx2 − 1 kb and − 0.2 kb promoter regions in young and aged BMSCs. The results were normalized to the percentage of various histone modifications in the input control. Data are shown as the mean ± SD. **p < 0.01, ***p < 0.001, aged versus young