Fig. 5

rCD248 interferes with melanoma vascular mimicry by binding to fibronectin. B16F10 cells treated with rCD248 proteins were subjected to A, B VM assay (network formation on Matrigel), C western blot analysis of MMP9 expression, and D cell proliferation assay (MTT assay). A Representative images of VM assay induced by the fibroblast-cultured conditioned medium (CM). B Statistical analysis of the total length of the network was measured 6 h after seeding on Matrigel. N = 3. *P < 0.05; ***P < 0.001 compared with vehicle control. C Western blot analysis of MMP9 expression in cells treated with CM in the presence of rCD248D1-4 (1 µM) for 1 day. N = 6. **P < 0.01. D B16F10 cells were cultured with or without rCD248D1-D4 for 1 to 3 days and followed by an MTT assay at indicated time point. N = 3. Soluble fibronectin reversed the inhibitory effects of rCD248 proteins on E, F cell migration and G, H VM. Soluble fibronectin (0.5 µM) was pre-incubated with rCD248D1-4 (0.5 µM) proteins for 30 min before the indicated experiments. (E) Representative images of the Boyden chamber migration assay using CM as a chemoattractant. F Statistical analysis of cell migration toward CM for 3 h. N = 4. **P < 0.01; ***P < 0.001. G Representative images of the VM assay. H Statistical analysis of the VM assay induced by CM. N = 4. ***P < 0.001