FIGURE 1.

Hh signaling activates OPN in RAW264.7 cells. A, RAW264.7 cells were treated with the indicated concentrations of recombinant SHH. The levels of OPN were assessed by real-time quantitative RT-PCR. Relative to untreated cells, the cells treated with SHH expressed significantly greater levels of OPN mRNA (p < 0.0001). B, OPN promoter construct (200 ng) and the β-galactosidase plasmid (200 ng) were co-transfected into RAW264.7 cells. Luciferase activity was assayed 24 h after SHH treatment and normalized to β-galactosidase. Each group was assessed in triplicate. The data are depicted as relative luciferase activity and are representative of three independent experiments. The increase in OPN promoter activity is significantly (p < 0.05) higher for the indicated groups relative to the control (untreated) cells. C, SHH treatment of the RAW264.7 cells (at the nm concentrations indicated) stimulates the expression of OPN. Cells were lysed 24 h after SHH treatment, and OPN and β-actin were assessed by immunoblotting. D, densitometric analyses of the immunoblotting results. The results are represented as band intensity relative to respective loading control. Band intensities are represented as arbitrary units.