Fig. 2

AICAR enhanced the BMP-2-dependent transcriptional activity on Ti via the expression of osteoblast differentiation-related molecules. A Id promoter site was fused to luciferase and transfected into MC3T3-E1 cells using Lipofectamine 3000. The cells were treated with BMP-2 (50 ng/mL) with or without AICAR (500 μM) for the indicated times during culture, and luciferase activity was assessed as a measure of Id transcriptional activity. The data are the mean from four culture wells (mean ± SEM). **Indicates P < 0.01 vs. treatment with BMP-2 alone. B–D MC3T3-E1 cells were incubated with BMP-2 in the presence or absence of AICAR (500 μM). Expression of the osteoblast differentiation-related genes Runx2 (Runx2), Osx/Sp7 (OSX/Sp7), and Ocn (Osteocalcin) in MC3T3-E1 cells after incubation with BMP-2 (50 ng/mL). Samples were analyzed by quantitative reverse transcription polymerase chain reaction and were normalized to the ß-actin mRNA. The data are the mean from six culture wells (mean ± SEM). **Indicates P < 0.01 vs. treatment with BMP-2 alone. E Cells were treated with BMP-2 (50 ng/mL) in the presence or absence of AICAR (500 μM). Western blotting was conducted using targeted and ß-actin antibodies. Similar results were obtained in four independent experiments