Fig. 5
Effect of SIRT1 activation/inhibition on Wnt signaling pathways during commitment of MSCs. 50 μM resveratrol or 10 mM nicotinamide was added to C3H10T1/2 cells with 20 μM 5-AZA during commitment for 3 days. a The relative expression profiles of CyclinD1 mRNA was determined using Realtime-PCR. b SIRT1, β-catenin and Cyclin D1 protein expression detected by western blotting at 24 h. c The luciferase reporter activity was measured at 24 h after transfection with resveratrol or nicotinamide. d RNAi SIRT1 plasmid was co-transfected into C3H10T1/2 cells with TCF-receptor. e Overexpression SIRT1 plasmid was co-transfected into C3H10T1/2 cells with TCF-receptor. The luciferase reporter activity was measured 48 h after transfection. f sFRP2 mRNA measured by Realtime-PCR and g Dact1 mRNA measured by Realtime-PCR at 0, 12, 24, 48, and 72 h. h Cells were fixed and stained with Oil Red-O on day 14 days after GM culture. i The quantification of h. The Oil-Red O stain was extracted with 100 % isopropanol and the optical density at 500 nm (OD500) was determined. j The relative expression profile of adipogenic marker genes: PPARγ, adiponentin were determined using Real-time PCR. k Cells were fixed and stained with Dil on day 14 days after GM culture. l The relative expression profile of myogenic marker genes: MyoD and MyHc were determined using Real-time PCR. Data represent mean ± SEM from three independent experiments. *P < 0.05; **P < 0.01