Figure 2.
Effects of FH535 treatment in the CaCo2 and HCT116 CRC cell lines. (a) FH5353 causes a dose- and time-dependent inhibition of cell proliferation. Cells were treated with the indicated concentrations of FH535 and counted after 24, 48, 72 and 96 h (x axis). Number of cell expansions are reported on y axis as mean values obtained from three independent experiments. (b) FH535 induces upregulation of μ-protocadherin (MUCDHL), E-cadherin (CDH1), p21waf1 and CDX2 transcripts. Cells were treated with the indicated doses of FH5353 and analyzed by QRT-PCR after 96 h for CaCo2 and 48 h for HCT116 cells. Mean variations of mRNA expression levels, deriving from three independent experiments, are reported in the y axis as relative fold-change (relative quantity). Results are represented as mean±S.E.M. values and asterisks indicate statistically significant results (P<0.05). (c) Western blot analysis following treatment with FH535 shows a decreased expression of TCF4 protein and a concomitant increase of MUCDHL and CDX2 protein expression. Cells were treated with the indicated doses of FH535 and analyzed after 96 h for CaCo2 and 48 h for HCT116 cells. This analysis was performed on nuclear and cytoplasmic extracts and normalized with actin. Analyzed proteins and their molecular weights are indicated on the left and on the right, respectively