FIGURE 4.

Activation of β-catenin signaling by TGF-β1 mediates PAI-1 induction. A, TGF-β1 induces β-catenin activation in tubular epithelial cells. HKC-8 cells were treated with TGF-β1 (2 ng/ml) for various periods of time as indicated, and the cell lysates were immunoblotted with antibodies against active β-catenin, total β-catenin, and GAPDH, respectively. B and C, ectopic expression of stabilized β-catenin promotes PAI-1 induction at basal and TGF-β1-stimulated conditions. HKC-8 cells stably transfected with either empty vector (pcDNA3) or FLAG-tagged, N-terminal truncated β-catenin (pDel-β-cat) were incubated in the absence or presence of TGF-β1 (2 ng/ml) for 48 h. A representative Western blot (B) and the quantitative data (C) are presented. CTL, control. **, p < 0.01 versus pcDNA3 controls. ††, p < 0.01 versus without TGF-β1 (n = 3). D–G, expression of stabilized β-catenin also promotes PAI-1 mRNA expression. HKC-8 cells stably transfected with either empty vector (pcDNA3) or FLAG-tagged, N-terminal truncated β-catenin (pDel-β-cat) were incubated in the absence or presence of TGF-β1 for various periods of time (D and E) at different concentrations (F and G). The quantitative data on PAI-1 mRNA induction (fold over the controls) are presented in E and G. *, p < 0.05 versus pcDNA3 controls. H, inhibition of β-catenin signaling blocked TGF-β1-induced PAI-1 expression. HKC-8 cells were pretreated with a specific small molecule β-catenin inhibitor (ICG-001) (10 μm) for 30 min, followed by incubation in the absence or presence of TGF-β1 (2 ng/ml) for 24 h.