HOXB9 expression and co-culture systems. (a) Cancer cells were co-cultured with HUVECs and human fibroblasts for 48 hours. Cytokines in supernatants were evaluated by multiplex assays (n = 24, each). (b) IL6 secretion levels in supernatants were compared by ELISA between the monoculture and co-culture systems in HT29-T or HCT116-T cancer cells. HOXB9 overexpression was induced by tetracycline (1 μg/ml). (c) VEGF secretion and bevacizumab treatment in the co-culture system, detected by ELISA (HT 29-T, left panel; WiDr-S right panel). (d, e) Tumor proliferation of HT29-T (d) and HCT116-T (e) cell lines on treatment with bevacizumab and/or an IL6-neutralizing antibody in the co-culture system with HUVECs and human fibroblasts. (f) IL6 secretion under treatment with bevacizumab and/or IL6 neutralizing antibody in the co-culture system with HUVECs and human fibroblasts. Error bars are SDs. ☆ p < 0.05 and † p < 0.005 by the U-test.