GSK-3 is inhibited during DC activation. MoDCs were prepared as in Figure 2. Cells were either continued in GM-CSF and IL-4 (iDC), or activated with E coli (1:1) or a baby hamster kidney cell line transfected with CD40L (BHK-CD40L, in a 20:1 ratio of DCs to BHK cells) in presence or absence of wortmannin (WM; 100 ng/mL). Cells were harvested and resuspended in lysis buffer 2 hours following activation. (A) Ser21/9 and Tyr279/216 phosphorylation of GSK-3 and Ser473 phosphorylation of Akt-1 were analyzed by Western blot. p38K analysis acts as protein loading control. One representative of 4 experiments is shown. (B) Strength of Ser9 phosphorylation of GSK-3β and (C) intracellular β-catenin expression were assessed by quantitative immunoblotting. Shown are the means ± SD; n = 4.