Fig. 5: CXCR7 contributes to melanoma angiogenesis and promotes VEGF secretion by upregulating HIF-1α expression.

a Immunofluorescent staining of CD31 (green) and DAPI (blue) in tumors derived from murine melanoma cells. The blood vessel densities were calculated by dividing CD31 area by DAPI area and were normalized to F0 Vec tumors (top) or F10 WT tumors (bottom). Left: representative images; scale bar = 50 μm. Right: relative quantitative results (n = 6–8 for each group). b Immunohistochemistry staining of VEGF in tumors derived from mice bearing murine melanoma cells. The staining intensities of VEGF were normalized to F0 Vec tumors (top) or F10 WT tumors (bottom). Left: representative images; scale bar = 100 μm. Right: relative quantitative results of staining intensities (n = 6–8 for each group). c The secretion of VEGF by murine melanoma cells with or without CoCl₂ (200 μM) treatment. Cells were pretreated with CoCl₂ for 6 h, and then incubated in serum-free medium containing CoCl₂ overnight. The conditioned mediums were collected and the levels of VEGF were determined by ELISA. d Immunohistochemistry staining of HIF-1α in tumors derived from mice bearing murine melanoma cells. The staining intensities of HIF-1α were normalized to F0 Vec tumors (top) or F10 WT tumors (bottom). Left: representative images; scale bar = 100 μm. Right: relative quantitative results of staining intensities (n = 6–8 for each group). e HIF-1α expression in murine melanoma cells treated with CoCl₂. f HIF-1α expression in melanoma cells under normoxic conditions. g A375 WT and KO cells transfected with siRNA targeting scramble control or HIF-1α were treated by CoCl₂. The conditioned mediums and cell lysates were collected to detect VEGF secretion and HIF-1α expression. ELISA assays were independently conducted in triplicate. The quantification of immunofluorescent and immunohistochemistry staining were evaluated in 12 random fields for each tumor. Data are presented as mean ± SD, *p < 0.05, **p < 0.01, ***p < 0.001; ns not significant