Figure 6. Smurf1 plays an important role in miR-17-mediated osteogenic differentiation of BMMSCs. BMMSCs were lentivirally transduced to upregulate the expression level of P53 (= pLenti-P53) or were transduced as lentiviral control (= pLenti-Cont). miR-17 was stable upregulated in BMMSCs by miR-17 mimics (= miR-17 mimics). miRNA control (= miR Cont). 16 Mon Cont = control BMMSCs, Smurf1 siRNA = downregulated Smurf1 level via si-RNA, miR-17 inhibitor = transfection with anti-miR-17. Statistically analyzed values show the mean ± SD (n=10). * p < 0.05. (A) Real-time PCR and western blot analysis on the expression of Smurf1 and TCF3 after upregulation of p53 (pLenti-p53) in BMMSCs derived from young mice. Normalization to ß-actin. (B) Western blot analysis on the expression of Smurf1. Transfection of miR-17 mimics in stable upregulated p53 BMMSCs derived from young mice. Normalization to ß-actin. (C-D) Real-time PCR and western blot analysis of Smurf1 expression in osteogenically differentiated BMMSCs from young and old mice. Normalization to ß-actin. (E) Alizarin red staining after osteogenic induction for 14 d of BMMSCs derived from old mice with/without siRNA-downregulated Smurf1 level and with/without transfection with miR-17 inhibitor. (F) Western blot analysis on old BMMSCs with/without siRNA-down-regulated Smurf1 level and with/without transfection with anti-miR-17 for the osteogenic markers Runx2, ALP, osterix. Normalization to ß-actin.