Figure 3: Ovatodiolide inhibits RCC cell migration, invasion, and tumorigenicity. (a) Transwell assay of cell migratory ability (upper) and invasive ability (lower) in four RCC cell lines. Five random fields of each transwell membrane were counted at 100x magnification. Data are mean SD of triplicate independent repeats. (b) Colony-forming assay of in vitro tumorigenicity. Data are mean SD of triplicate experiments. (c) Effect of ovatodiolide (40 μM) and cyclohexamine (CHX, 100 μg/mL) or MG-132 (10 or 20 μM) for 48 hr. (d) RCC cells were treated with doses of ovatodiolide or DMSO for 48 hr. Western blot analysis of phosphorylation of AKT (p-AKT [S473]) and inhibitory regulation of p-GSK-3β (S9) and β-catenin (p-β-catenin [S552]) and inactive form of β-catenin (phosphorylation at S33/37, T41) in four RCC cells.