Fig. 2. Pyroptosis of endothelial cells in the aorta of nicotine-induced ApoE^−/− mice.

a Comparison of expression and subcellular distribution of caspase-1 by double staining of caspase-1 (green) and CD31 (red) in atherosclerotic lesions of ApoE^−/− mice among various groups: ND, ND + Ni (nicotine), HFD, or HFD + Ni. Note the presence of caspase-1 in endothelial cells as indicated by the co-localization of caspase-1 and CD31 (an endothelial marker). Magnification: ×200. b Identification of endothelial cell death by co-localized staining of CD31 (red) and TUNEL (green). The nuclei were stained blue with DAPI. Scale bar = 100 μm. Magnification: ×200. c Increases in the protein levels of caspase-1 by nicotine in the HFD group, as revealed by western blot analysis. d–h Increases in the expression of pyroptosis-related genes (NLRP3, ASC, caspase-1, IL-1β, and IL-18) at both protein and mRNA levels by nicotine in intimal samples of ApoE^−/− mice of the HFD group. n = 6 mice for each group. The data are presented as the mean ± SEM., *P < 0.05. i–j Elevation of mean serum concentrations of IL-1β and IL-18 by nicotine in HFD-fed mice, as determined by ELISA assay. The data are shown as mean ± SEM. n = 6 mice in each group. *P < 0.05