Microenvironment Promotes Tumor Cell Reprogramming in Human Breast Cancer Cell Lines
Figure 5
Cdh1 gene expression and its regulation.
(A) Real-time PCR for Cdh1 gene in MDA-MB-231 cells treated for 5 days with EW. The results are the mean (SEM) of six independent experiments; *p<0.05 vs ctrl by Mann Whitney test. (B) Methylation of the Cdh1 gene promoter in MDA-MB-231 cells at 3 and 5 days of treatment with EW. Visible PCR product in lanes U indicates the presence of unmethylated sequences; visible PCR product in lanes M indicates the presence of methylated sequences. (C) and (D), representative western blot showing the levels of ZEB1 and ZEB2 proteins in MDA-MB-231 cells after 3 and 5 days of treatment with EW; proteins signals are shown together with α-tubulin bands, chosen as loading control. (E) Representative western blot showing the levels, after 3 and 5 days of treatment with EW, of the histone H3 trimethylated at lysine 4 (H3K4me3), and the levels of the total histone H3 (H3 tot) in MDA-MB-231 cells. Below is shown the densitometric quantification of H3K4me3, obtained normalizing the O.D. of protein bands versus the O.D. of Histone H3. (F) Representative western blot showing the levels, after 3 and 5 days of treatment with EW, of LSD1 and SNAIL proteins in MDA-MB-231 cells. Below is shown the LSD1/SNAIL ratio, obtained normalizing the O.D. of LSD1 bands versus the O.D. of SNAIL. The results are the mean (SD) of three independent experiments; *p<0.05 vs ctrl by Student’s t test.