Fig. 5: Elucidation of the mechanism by which CAFs increased ERβ expression in BCa cells.

a IGF-1 levels in the CM from CAFs, BCa cells or co-cultured (CAF+BCa) cells were quantified by ELISA (left panel). IGF-1 levels in the CM of co-cultured cells treated with different concentrations of cisplatin for 48 h were quantified by ELISA (right panel). b Assessment of IGF-1 mRNA expression in CAFs by qRT-PCR using the β-actin gene as the normalization control. c Immunofluorescence double staining for α-SMA (green) and IGF-1 (red) in pathologic non-response RC tissues (400×). Double-positive areas are indicated (arrows). d Western blotting (left panel) and qRT-PCR (right panel) results showed that blocking IGF-1 with an anti-IGF-1 neutralizing antibody can partially reverse CAF-mediated ERβ and Bcl-2 upregulation in BCa cells. The mean expression value of the control cells (T24, 5637) was defined as 1. e Blocking IGF-1 can partially reverse the effects of CAFs on BCa cell resistance to apoptosis induced by cisplatin. f Western blotting (left) and immunofluorescence staining (right) show that blocking IGF-1 in the co-culture system reversed the capacity of BCa cells to induce NF transformation into CAFs (The expression of α-SMA in the fibroblasts decreased after blocking IGF-1 in the co-culture system). g Blocking IGF-1 in the co-culture system reversed the capacity of the BCa to recruit fibroblasts. h IGF-1 increased the protein (left panel) and mRNA (right panel) levels of ERβ and Bcl-2 in BCa cells. i IGF-1 can increase BCa cell resistance to cisplatin. The procedures in d, e, h and i were conducted after the cells were incubated in an appropriate concentration of cisplatin (T24, 25 mg/L; 5637, 6 mg/L). Data are presented as the mean ± SD. *P < 0.05