Figure 8.

Differential β-catenin immunoreactivity by MSCs cultured under various conditions. MSCs were fluorescently labeled with β-catenin antibody (green) and DAPI nuclear stain (blue), after culturing cells for 2 days in the absence or presence of BIX01294 and/or TSA and for an additional 2 days with or without Wnt11 or Wnt3a. (a) MSCs cultured in sequence with BIX01294 and Wnt11 were deficient in cells that brightly stained for β-catenin. In contrast, (b) cultures treated sequentially with TSA and Wnt11 still displayed numerous brightly stained β-catenin-positive cells. (c) However, MSCs pretreated with both BIX01294 and TSA, followed by Wnt11 exposure, no longer displayed cells with high-intensity β-catenin immunoreactivity. (d) MSCs treated with Wnt3a showed a marked increase in β-catenin-positive cells. (e) Yet, this high-intensity β-catenin immunoreactivity disappeared when MSCs were first exposed to BIX01294 prior to treatment with Wnt3a. Scale bar = 50 μm. (f) Tabulation of immunofluorescent cells under these various conditions verified that BIX01294 pretreatments decreased the number of brightly stained β-catenin-positive cells that were derived from the cultures. Statistical significance is indicated by ^∗∗p < 0.01.