Fig. 4.

SelS is a major regulator of NET formation mediated by Se-deficient arteritis. (A)–(B) Heatmap (A) and volcano plot (B) of 25 selenoprotein mRNA expression levels in broiler aortas. (C) The protein expression levels of SelS in broiler aortas (N = 3; Bars that do not share the same letters are significantly different (P < 0.05) from each other). (D) The protein expression levels of SelS in PAECs cocultured with or without PMA-stimulated and with or without DNase Ⅰ-treated neutrophils (N = 3, ***P < 0.001, compared with the normal Neutrophils + PAECs; ^###P < 0.001, compared with the PAECs cocultured with PMA-stimulated but without DNase Ⅰ-treated neutrophils). (E) After transfection with siSelS for 24 h, the protein expression of SelS in PAECs. (F) LDH release of PAECs after transfection with siSelS for 24 h (N = 3, ***P < 0.001). (G) IF analyses of TNF-α (red) and IL-1β (green) in PAECs after transfection with siSelS for 24 h (scale bar, 20 μm). Fields from one representative experiment of three are shown. (H) Quantification of relative TNF-α and IL-1β intensity in PAECs after transfection with siSelS for 24 h (N = 3, ***P < 0.001, compared with the NC group for TNF-α; ^###P < 0.001, compared with the NC group for IL-1β). (I) Fluorescence microscope for NETs (green) of neutrophils cocultured with PAECs transfected with or without siSelS (scale bar, 50 μm). Fields from one representative experiment of three are shown. (J) Quantification of NET formation by fluorescence microscope in neutrophils co-cultured with PAECs transfected with or without siSelS (N = 3; ***P < 0.001). Results are presented as mean ± SEM. Statistical significance was obtained by one-way ANOVA or unpaired Student's t-test. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)