Figure 4

TGFβ-induced caspase-9 cleavage requires both Par6 and TβRI activation in NMuMG 3D structures. 12-day old 3D structures of the indicated NMuMG-derived cell lines were treated with control media (DMSO alone), TGFβ1 (TGFβ), the TβRI/Smad2 inhibitor SB-431542 (SB) or TGFβ + SB for 48 hours, fixed and immunostained for zonula occludens-1 (ZO-1, green) and cleaved caspase-9 (CC9, red). Dapi was used to visualize the nuclei. A. Confocal images show representative structures for each given cell line and treatment. Scale bar = 20 μm. Arrow = apoptotic nuclei. B. Average percentage of apoptotic structures was calculated per each cell type and treatment, with n = 3 for biological replicates. Two-way ANOVA for all cell lines and treatments was significant (p < 0.001). Bonferroni post test compared differences between control and treatment for each cell line, **p < 0.001.