Fig. 4: Epithelial to mesenchymal transition (EMT) is a hallmark of NETosis in PDAC tumours.

A Gene set enrichment analysis (GSEA) of differentially expressed genes in group 2 (poor outcome) versus group 3 (better outcome). Bar plot showing pathways upregulated in patient group 2 (poor prognosis) versus patient group 3 (better prognosis). B Analysis of Hallmark epithelial-mesenchymal transition (EMT) genes significantly upregulated in patient group 2 compared to patient group 3. Grey bars, strength of significance of gene dysregulation; Red squares, magnitude of differential expression; Teal bubbles, difference in protein levels for each gene; Brown ribbons, gene-gene relatedness based on GeneFriends analysis. C Western blot analysis of the indicated EMT markers in MIA PaCa-2 cells cultured in the presence of increasing concentrations of NETs for 24 h. Western blot analysis of NET-induced EMT markers in response to dox-inducible knockdown of ILK expression in D MIA PaCa-2 cells cultured with 20 µg/mL NETs for 24 h and E KPCY cells cultured with 20 µg/mL NETs for 48 h. Western blot analysis of NET-induced Zinc-finger E-box-binding homeobox 1 (ZEB1) expression in response to pharmacologic inhibition of ILK in F MIA PaCa-2 cells and G KPCY cells cultured with 10 µg/mL NETs for 48 h in the presence of 10 µM of ILKi. H Western blot analysis of NET-induced ZEB1 expression in MIA PaCa-2 cells cultured on 2.5% Matrigel with 10 µg/mL NETs for 48 h and exposed to 10 µM of the ILK inhibitor. I Western blot analysis of NET-induced ZEB1 expression in MIA PaCa-2 cultured with 10 µg/mL NETs for 48 h on the indicated substrates in the presence of 5 µg/mL function-blocking Ab against ITGB1. Quantification of band intensities is reported below the blots (C–I).