Fig. 5

Effect of CXCL12, co-culture with colorectal cancer cells and IL-1Ra pretreatment on angiogenesis. (A) Effect of CXCL12 on angiogenesis. After incubation of HUVECs/fibroblasts in the presence or absence of CXCL12 or anti CXCL12 Ab for 11 d, angiogenesis were stained with CD31 antibody. (a: Control; a-1: Culture system treated with 1 ng/mL CXCL12; a-2: Culture system treated with 10 ng/mL CXCL12; a-3: Culture system treated with 100 ng/mL CXCL12; a-4: Culture system reated with 100 ng/mL CXCL12 and 1 μg/mL of anti CXCL12 Ab (Magnification: × 40). (B) Effect of co-culture with different metastatic potential colorectal cancer cells (HT-29) and (CaCo-2) on HUVEC tubular formation. HUVECs/fibroblasts were co-cultured with HT-29 or CaCo-2. Theco-cultured system was incubated for 11 days, and the tube formation was measured as described earlier. Magnification: × 200. (b: Control; b-1: co-cultured with HT-29 cells; b-2: c Co-cultured with CaCo-2 cells). (C) Effect of co-culture with HT-29 and CaCo-2 cells pretreated with CXCL12, IL-1Ra and CXCR4 siRNA on on HUVEC tubular formation. (c: co-cultured with HT-29 cells; c-1: co-cultured with HT-29 cells pretreated with 10 ng/mL CXCL12; c-2: co-cultured with HT-29 cells pretreated with 10 ng/mL of IL-1Ra; c-3: co-cultured with HT-29 cells tranfected with CXCR4 siRNA duplex oligoribonucleotide; c-4: co-cultured with CaCo-2 cells; c-5: co-cultured with CaCo-2 cells pretreated with 10 ng/mL CXCL12; c-6: co-cultured with CaCo-2 cells pretreated with 10 ng/mL of IL-1Ra; c-7: co-cultured with CaCo-2 cells tranfected with CXCR4 siRNA duplex oligoribonucleotide). Columns represent mean pixels of HUVEC tube formation area; error bars represent SD. Multiple comparisons were made by using one-way ANOVA, followed by Student–Newman–Keuls test. *P < 0.01