Figure 4

Hey2 is upstream of Sox17 and RhoU/Wrch1 is upstream of both Hey2 and Sox17. (a) SBEC cholangiocytes transfected with RhoU/Wrch1 or Hey2 plasmids and with an empty vector as a control. Cells were immunostained for RhoU/Wrch1 or Hey2 (red) 48 h after transfection. Nuclei were stained with DAPI (blue). Images are representative of n = 3 independent experiments. Relative fluorescence intensity was calculated from at least 20 fields for each condition. (b) SBECs transfected with RhoU/Wrch1 or Hey2 plasmids. RhoU/Wrch1 and Hey2 mRNA levels were evaluated by qRT-PCR 48 h after transfection. All graphs represent mean ± s.e.m, n = 3-7 independent experiments. NS = non-significant. (c) SBECs were treated with siRNA against RhoU/Wrch1 or with scrambled RNA for 48 h. After 24 h, cells were treated with biliatresone (2 µg/ml) or DMSO (same volume as biliatresone) for an additional 24 h. RhoU/Wrch1, Hey2, and Sox17 transcript levels were evaluated by qRT-PCR. Black histograms highlight biliatresone treatment; n = 4-10 independent experiments. All graphs represent mean ± s.e.m.