Figure 5. Long-term intermittent dosing with RAP suppresses markers of proliferation and increases differentiation after 7 weeks of treatment (N = 5).

(A) and (B) Immunostaining of mouse distal colon shows increased phospho-mTOR (p-mTOR) and phospho-S6 (p-S6) expression in mouse polyps from CDX2P-CreER^T2 Apc^fl/fl mice injected with TAM. Both increases are suppressed by RAP treatment. (C) BrdU immunostaining showed increased incorporation of BrdU in mouse polyps and RAP partially reversed the effect. (D) Alcian blue staining shows loss of most of the differentiated goblet cells in mouse polyps and RAP restores the goblet cell differentiation. (E) Immunostaining shows increased expression of Sox9 (a β-catenin-regulated cell fate marker) expression in polyps and RAP decreased the expression. (F) Immunostaining shows increased nuclear and cytoplastic β-catenin staining in polyps and RAP treatment restores the membrane staining in most of the cells except a few strongly staining crypts. Mice assessed in the RAP group when moribund had rare areas of polyps with HGD that generally stained like mice in the vehicle group for all markers assessed (column 4, A–F). Scale bars  =  100 µm. The inserts are at two times higher magnification than the original.