Inhibition of Akt and COX-2 expression by embelin in PC3 cells.

<p>(A) Cells were treated with 30 μM embelin for the indicated time periods, and whole cell lysates were prepared, and extracted proteins were resolved by SDS-PAGE (10%) and Western blot analysis using phospho-Akt, total Akt, and COX-2 antibodies was conducted. The numbers between the blots are the ratios of the intensity of bands after normalized with control. (B) Cells were transfected with Myr-Akt plasmid and then treated with 30 μM of embelin for 24h. Cell viability was measured by CCK. Values represent mean ± S.D. of three independent determinations. <b>*,</b> significantly different from the untreated control cells (<i>p</i>< 0.01) and **, significantly different from the embelin only-treated cells (<i>p</i><0.001). Whole cell lysates were prepared and the levels of total Akt and phospho-Akt were determined by Western blot analysis. (C) AKT inhibitor IV treated cell lysates were prepared and extracted proteins were resolved by SDS-PAGE (10%) and Western blot analysis using phosphor-Akt, total Akt, and GAPDH antibodies was conducted. Cells were treated with 0.312 μM of AKT inhibitor IV for 24 h. Cell viability was measured by CCK. Formazan formation was quantified by spectrophotometry at 450 nm. The percentage of cells surviving in each group relative to the control was calculated. (D) Cells were transfected with COX-2 luciferase vector and renilla luciferase vector for 48 h. Cells were subjected to the dual-luciferase assay. The relative firefly luciferase activity, normalized by the renilla luciferase activity, is shown. Values represent mean ± S.D. of three independent determinations.<b>*,</b> significantly different from the untreated control (<i>p</i>< 0.05).