4-MU treatment inhibited tumour growth of OSC1 cells in nude mice. Nude mice received subcutaneous injection of 106 OSC1 cells, and tumour growth was monitored for 47 consecutive days. (A) Tumour volume as determined by calliper measurements; mean ± SEM, n = 21 (control), 17 (4MU); *, p < 0.05. (B) Representative fpVCT images of control mice and 4-MU treated mice. Scale bars, 1 cm. Average tumour volumes after sacrification of mice at the end of the experiment as measured by fpVCT showed a significant reduction in tumour volume in the 4-MU group. (C) Cryosections stained with H&E demonstrated a differentiated tumour phenotype in mice treated with 4-MU. This phenotype was characterised by larger tumour cell clusters with a cell-rich border region (arrows) and pronounced tumour stroma. Scale bars, 200 μm. (D) Stromal fibroblasts were stained with alpha-smooth muscle actin staining (red). Scale bars, 200 μm. (E) Hyaluronan staining (red) was associated with both tumour cells and stromal tissue and was markedly reduced by treatment with 4-MU. Scale bars, 200 μm. Shown are representative images of n = 6 (control) and n = 7 (shHAS3) experiments.