JCS019455 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure S1 (Adobe PDF) -

  Fig. S1. The JNK inhibitor SP600125 inhibited TGF-β1-induced expression of mesenchymal genes. MTEC cells were treated with 5 ng/ml TGF-β1 in the presence or absence of SP600125 (10 µM) for 3 days. TGF-β1-induced mesenchymal gene expression was evaluated by Taqman analysis. Results are expressed as relative expression compared to sham controls (* P<0.05 compared to TGF-β1 groups, ANOVA).

• Supplemental Figure S2 (Adobe PDF) -

  Fig. S2. Inhibition of JNK1 by SiRNA-knockdown-attenuated TGF-β1-induced expression of PAI-1. C10 cells transfected with control or JNK1-specific SiRNAs (100 nM) were exposed to 5 ng/ml TGF-β1 for 2 days. TGF-β1-induced PAI-1 gene expression was evaluated by Taqman analysis. Results are expressed as relative expression compared to sham controls (* P<0.05 compared to the TGF-β1 group, ANOVA). Inset: confirmation of loss of JNK1 protein content but not JNK2 following exposure of cells to JNK1 SiRNA. Expression of total JNK was evaluated by western blotting. β-Actin represents loading control.

• Supplemental Figure S3 (Adobe PDF) -

  Fig. S3. The generic caspase inhibitor zVAD did not alter TGF-β1-induced loss of TER. MTEC cells were treated with 5 ng/ml TGF-β1 in the presence or absence of zVAD (10 µM) for a total of 10 days. TER was measured daily. Results are expressed as mean±s.e.m. (n=3).

• Supplemental Figure S4 (Adobe PDF) -

  Fig. S4. The generic caspase inhibitor zVAD did not affect TGF-β1-induced EMT in vitro. MTEC cells were treated with 5 ng/ml TGF-β1 in the presence or absence of zVAD (10 µM) for 6 days. TGF-β1-induced mesenchymal gene expression was evaluated by Taqman analysis and the results expressed as relative expression compared to sham controls (* P<0.05 compared to sham and z-VAD groups, ANOVA).

• Supplemental Figure S5 (Adobe PDF) -

  Fig. S5. The generic caspase inhibitor zVAD prevented FasL-induced cleavage of caspase 3 in MTEC cells. MTEC cells were incubated with zVAD-FMK (10 µM) or DMSO for 2 hours before treatment with FLAG-FasL (500 ng/ml) plus cross-linking antibody, anti-FLAG M2 antibody (2 µg/ml) (FasL) or M2 as a reagent control (control) for 6 hours. Caspase 3 cleavage was evaluated by western blotting using an antibody specific for cleaved caspase-3. β-Actin represents loading control. Samples were run in duplicate.