The synergy of Wnt and AR promotes proliferation in prostate cancer cells. (A) PC3 cells in phenol-red free RPMI with 5% charcoal-stripped serum were transfected with the indicated plasmids and treatments in 96 well plates. 7 days after transfection, the number of cells was determined with a Guava proliferation assay. Cells transfected with GFP were used as control. The percentage of cell numbers was calculated for cells transfected with Wnt or AR or both compared with control samples. (B) AR expression and Flag-tagged AR in LNCaP-Flag-AR cells and LNCaP cells were detected by Western blot. Actin was used as loading control. (C). LNCaP cells and LNCaP-Flag-AR cells in phenol-red free RPMI with 5% charcoal-stripped serum were treated with 100 ng/ml Wnt3A for four days in 96 well plates. The cells were then labelled with [3H]-thymidine and harvested to measure the amount of thymidine incorporation. LNCaP cells which received no Wnt3A treatment were used as control. The percentage of [3H]-thymidine incorporation was calculated compared with control.