Curcumin, 5-FU and the combinational treatment suppress TGF-β-mediated synergistic crosstalk between CRC-cells and fibroblasts in tumor microenvironment co-cultures.

<p>A-B: HCT116 high density mono-cultures were either left untreated (HCT, Co.) or were co-cultured with MRC-5 in monolayer. Tumor microenvironment co-cultures were either left untreated (Co.), treated with curcumin alone (5µM), 5-FU alone (1, 5, and 10µM) or were pretreated for 4 h with curcumin (5µM) followed by treatment with 5-FU (0.1, 1, 2, 3µM). After 10 days of culture, total cell lysates of HCT116 high density cultures were prepared and immunoblotting performed for TGF-β3 (A) or p-Smad2 (B). C-D: HCT116 high density mono-cultures were either left untreated (HCT, Co.) or were co-cultured with MRC-5 in monolayer. Tumor microenvironment co-cultures were either left untreated (Co.) or treated with neutralizing pan-TGF-β antibody (10, 20, 30 ng/ml). After 10 days of culture, total cell lysates of HCT116 high density cultures were prepared and immunoblotting performed for TGF-β3 (C) or p-Smad2 (D). Densitometric evaluation of protein expression as revealed by western blot analysis was performed in triplicate. Housekeeping protein β-actin served as a loading control in all experiments. Values were compared to the control and statistically significant values with <i>p</i><0.05. Significant values are marked with (*).