FIGURE 6.

NETs promote EGFR‐dependent EMT in vivo. A. Ventral flanks of female BALB/c nude mice were injected with 2.5 × 10⁶ MIA PaCa2 cells subcutaneously. When the tumours reached approximately 4 × 4 mm in size, tumour‐bearing mice were randomly assigned to different experimental groups (n = 4 per group) and treated with different types of conditioned medium. We seeded neutrophils at a density of 5 × 10⁶/ml with or without PMA and collected the conditioned medium 8 h later. Collected conditioned medium (CM) was concentrated using an Amicon Ultra Centrifugal Filter device (Merck Millipore) with a molecular weight cut‐off of 10 kDa. 10 ml conditioned media were concentrated to 0.5 mL. Intratumoural injection of 500 µL concentrated CM was performed every day. The volume was calculated according to the formula: V = ½ × length × width². On day 24, the tumours were excised and photographed. The data were shown as mean with SD of four mice in each group. B, Immunohistochemistry staining of nude mice tumour tissue sections for EGFR, p‐EGFR, N‐cadherin and vimentin were shown. C, Immunohistochemistry staining of citH3, p‐EGFR E‐cadherin in PDAC patients. D, Relationship between CitH3, p‐EGFR and E‐cadherin expression in 150 PDAC patients’ tissues