Fig. 1.

Time-, dose-, and TGF-βRI kinase-dependent cytotoxicity of TGF-β in organoids with a mutant β-catenin/TCF pathway. (A) Immunostaining for active caspase-3 in ApcV∆/∆ organoids with or without TGF-β treatment (3 ng/mL for 16 h). Apoptotic cells in the outer epithelial layer of TGF-β–treated organoids are indicated with the dotted line. (B) Representative images and fold increase of active caspase-3 positivity in the TGF-β–treated ApcV∆/∆ organoids (flow cytometry measurements from three control or TGF-β–treated organoids). (C) Phase-contrast microscopic images of ApcV∆/∆ organoids with and without 48 h TGF-β treatment. (D) TGF-β dose–response curve in ApcV∆/∆ organoid cultures treated continuously for 48 or for 4 h, with analysis 44 h thereafter. (E) Effect of the TGF-βRI kinase inhibitor SB431542 (10 µM) in the cultures treated for 48 h. (F) The effect of the caspase inhibitor Z-VAD-FMK (50 μM) on the TGF-β–induced (1 h) death of ApcVΔ/Δ organoids, analyzed at 48 h. (G) Response of the indicated Apc-mutant organoids to a 4-h TGF-β treatment. The organoids isolated from the small intestine are labeled as “si” and those from colon as “co.” [Scale bars: 50 µm (A) or 100 µm (C).]