Figure 1

CAR modulation on the TNF-α/TGF-β1-induced EMT. U87MG cells were treated with TNF-α (10 ng/ml)/TGF-β1 (10 ng/ml) in the absence or the presence of CAR (10 µM) in complete medium for 48 h. (A) At the end of the incubation, representative images were taken. (B) Real Time RT-PCR analysis of the EMT markers. The data were the mean values from three different experiments. (C,D) U87MG cells were treated as described above, and the levels of the EMT markers were evaluated by Western blotting with the use of specific antibodies. One representative Western blot is presented (C). The bar graph (D) shows the densitometric analysis of the Western blot, which was performed using the ImageJ program. The data are presented as the means of three different experiments. (E) Real Time RT-PCR analysis of the transcription factors that act as master regulators of the EMT (Snail, Slug, Twist and ZEB1) (F) U87MG cells were treated as indicated, and the levels of miR-200c were quantified at the end of the incubation. The data are presented as the means of three different experiments each performed in duplicate. The significance of the differences was determined by one-way ANOVA, followed by Bonferroni’s post hoc test: *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001 vs. the control; ^#P ≤ 0.05, ^##P ≤ 0.01, ^###P ≤ 0.001 vs. TNF-α/TGF-β1 alone; ^§§P ≤ 0.01, ^§§§P ≤ 0.001 vs. CAR alone.