Fig. 6. Inhibition of Nampt promotes FoxO and β-catenin acetylation and decreases osteoblastogenesis.
a–e Bone marrow stromal cells from WT or Namptf/f mice, infected with Ad-Cre and cultured under osteogenic conditions for 5 days. a mRNA expression by qRT-PCR. b Relative cellular NAD+ levels. AFU arbitrary fluorescence units. c Protein levels by western blot. Each band represents one mouse. d mRNA expression by qRT-PCR. e Representative images (left) and quantification of Alizarin Red staining (right) of bone marrow stromal cells, described in a, cultured under osteogenic conditions for 21 days. f mRNA expression in bone marrow stromal cells from 3-month-old control (FoxO1,3,4f/f) or cKO (FoxO1,3,4ΔOsx1-Cre) littermate mice cultured under osteogenic conditions for 5 days. g Time-dependent effects of 10 µM FK866 on NAD levels in bone marrow cells from 6-month-old C57BL/6 female mice. h Representative images (top) and quantification of Alizarin Red staining (bottom) of bone marrow stromal cells described in f, cultured under osteogenic conditions without or with 10 µM FK866 for 21 days. In all experiments except c, data were obtained from triplicate cultures from cells pooled from three mice per group. Data represent mean ± SD; p values by two-tailed t-test (a–d, f), one-way ANOVA (g), or two-way ANOVA (h).