Fig. 4

CAFs-derived IL-6 enhances survival and promotes migration and invasion in RT4 bladder cancer cells. RT4 cells were cultured in the CM from HFs, HFs + TGFß or iCAFs with or without the anti-IL-6 antibody (1 μg/mL; x-IL-6). a. The proliferation of the RT4 cells was evaluated using the CyQUANT Kit. b. The migration of RT4 cells was determined by using Ibidi-silicone insert. Insert margins were marked by white vertical lines on optical micrographs. After 6 h and 12 h, the percentage of wound closure was evaluated by measuring migration distances (spaces between the white vertical lines). c. The invasion of RT4 cells was measured by transwell assays. Cells that had migrated through the membrane were fixed and nuclei were stained with DAPI. Representative photographs of migratory cells on the membrane are shown. The relative cell migration was determined by the number of migrated cells in 10 randomly selected fields. d. The sensitivity of RT4 cells to mitomycin C (0.5 μg/mL; Mito C) was evaluated using The CyQUANT Kit. RFU means relative fluorescence unit. All values are averages of three independent replicates. The graphs show mean +/− SD. The difference between groups was analyzed by one-way ANOVA followed by post hoc analysis using Dunnett’s multiple comparison tests. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, n.s. = non-significant, n = 5–25