Fig. 2

Immunofluorescent characterization of growing colonies. A–D Representative images depicting staining for surfactant protein c (Sftpc) in air-exposed (A), continuous cigarette smoke (CS)-exposed (B), and intermittent CS-exposed (C) mice. No significant difference was noted in the ratio of the number of Sftpc-positive cells in colonies among the three groups. Data are presented as means ± SD. N = 4–5 per group. Scale bars: 100 μm. E–H Representative images depicting staining for aquaporin-5 (Aqp5) in air-exposed (E), continuous CS-exposed (F), and intermittent CS-exposed mice (G). No significant difference was observed in the ratio of Aqp5-stained area per cell among the three groups (H). The ratio in each group was calculated based on the average value of the air-exposed group. Data are presented as means ± SD. N = 5–6 per group. Scale bars: 100 μm. I–K Colonies immunofluorescence-stained for E-cadherin and vimentin. Scale bars: 100 μm. L–N Representative images depicting staining for PCNA in air-exposed (L), continuous CS-exposed (M), and intermittent CS-exposed mice (N). Scale bars: 100 μm. O Comparison of PCNA positive cell ratio in colonies growing in 3D culture in air-exposed, continuous CS-exposed, and intermittent CS-exposed mice. Data are presented as means ± SD. N = 8 (air), 13 (continuous CS exposure), and 5 (intermittent CS exposure)