Fig. 3. Antibiotic-treated mice have a higher number of tumour foci and lower lung-infiltrating IL-9-producing T cells.

Antibiotic-treated (ABX) and -untreated (control) C57BL/6 mice were intravenously injected with 5 × 10⁴ B16F10 cells. Tumour foci number in the lungs was determined 14 days after injection (a). Lungs were then digested, and cells stimulated with PMA (50 ng/ml), ionomycin (500 ng/ml) and Brefeldin A (5 μg/ml) for 4 h at 37 °C and 5% CO₂. The frequency of IL-9-producing CD4 + (b) and CD8 + (c) T cells and IL-9-producing non-T cells (d) was evaluated by flow cytometry. Metagenomic analysis of the gut microbiota was performed in faecal samples collected at the day of and 2 weeks after injection of B16F10 cells (e). Mice from the ABX group were kept under treatment throughout the experiment. Data are shown as mean ± SD. Unpaired Student’s t test was used for statistical analysis.