Fig. 3.

CCN2-mediated activation of Akt/GSK-3β signaling pathway is sensitive to PI3-kinase- and Akt-inhibiton. a Western blot analyses of phosphoprotein and total protein levels of Akt, b-c GSK-3β, and d p70S6K in cardiac myocytes incubated for 15 min without (Control) or with 100 nmol/L rhCCN2 (CCN2), or with 100 nmol/L rhCCN2 in the presence of the PI3-kinase inhibitor LY294002 (CCN2 + LY) or in presence of the Akt inhibitor API-2 (CCN2 + API-2). e Inhibition of phospho-Akt (Ser473) activities by competitive inhibition of pleckstrin homology (PH) domain of Akt by expression of GRK2ct. Western blot demonstrating phospho-Akt (Ser473) and total Akt levels in cardiac myocytes expressing GRK2ct or LacZ in the absence or presence of rhCCN2. Cells were incubated for 40 h after infection with Ad-GRK2ct or Ad-LacZ at 1,000× multiplicity of infection (MOI), then stimulated with rhCCN2 (100 nmol/L) for 30 min. f Western blot analysis of GRK2ct expression in cardiac myocytes 40 h after infection with 1,000 MOI Ad-GRK2ct or Ad-LacZ. The histograms demonstrate densitometric analysis of indicated phospho-protein activities relative to total contents of the respective proteins. Data are mean ± SEM, n = 3 in each group, *p < 0.05. The figure is representative of three independent experiments