Fig. 4

Mettl14 knockout promoted bladder TIC self-renewal. a Mettl14 knockout cells were used for oncosphere formation. Two weeks later, typical images of spheres were taken and sphere-formation ratios were quantitated. b The ratios of CD44⁺ bladder TICs in Mettl14 knockout cells were analyzed by FACS. c Mettl14 knockout and control cells were used for Ki67 staining, and counterstained with DAPI. Typical images were shown in upper panel and Ki67⁺ cell ratios were shown in lower panel. d, e Mettl14 knockout and control cells were used for transwell invasion assay. 36 h later, invasive cells were stained with crystal violet for visualization. Typical images were shown in D and cell numbers were counted in e. f 5 × 10⁶ Mettl14 knockout cells were subcutaneously injected into BALB/c nude mice, and tumor weight was measured one month later. n = 6 for each sample. g, h 10, 1 × 10², 1 × 10³, 1 × 10⁴, and 1 × 10⁵ Mettl14 knockout and control cells were used for 3 months’ tumor initiation assay. Six BALB/c nude mice were used per sample and the ratios of tumor formation mice were shown (g). Bladder TIC ratios were analyzed by extreme limiting dilution analysis (h). VS, versus. Six patients were all examined for tumor initiation assay, and only the results of patient #1 were shown. i Mettl14 knockout T24 cells were used for oncosphere formation, and typical images of spheres were taken two weeks later. j Mettl14 knockout T24 cells were used for transwell invasion assay. 36 h later, invasive cells were stained with crystal violet for visualization. k Mettl14 knockout T24 cells were used for tumor propagation assay, and the picture of established tumors was taken. *P < 0.05, **P < 0.01, ***P < 0.001, by two-tailed T test. At least three independent experiments were performed and got similar results