Fig. 6. NLRP3-mediated endothelial cell pyroptosis in a KD mouse model.

Mice were continuously injected i.p. with 4 mg CAWS for 4 days to induce coronary arteritis. Twenty-eight days later, mice were sacrificed and the heart tissues were harvested for follow-up examinations. a Representative images from H&E staining at 28 day post-CAWS injection. Scale bar = 100 μm. b Expression of macrophage marker F4/80 was determined using IHC staining in the endothelium of coronary arteries. The histogram showed the area percentage of the endothelium positive for F4/80 in coronary arteries. Enlarged images of area of interesting (AOI) were indicated with a red arrow. Scale bar = 100 μm. c The expression of neutrophil marker was examined in the endothelium of coronary arteries. The histogram exhibited the area percentage of the endothelium positive for neutrophil marker in coronary arteries. Scale bar = 100 μm. d Expression of pyroptosis-related proteins was examined by western blot analysis. e–k Quantitative analysis of pyroptosis-related protein expression was conducted. Data were expressed as mean ± SD (n = 3). l Comparison of expression and subcellular distribution of caspase-1 by double staining of caspase-1 (green) and CD31 (red) in coronary artery of control mice, CAWS mice, and CAWS mice pretreatment with MCC950. The presence of caspase-1 in endothelial cells was indicated by the co-localization of caspase-1 and CD31 (an endothelial marker). Magnification: ×200. Scale bar = 50 μm. m Identification of DNA fragmentation by co-localized staining of TUNEL (green) and CD31 (red). The nuclei were stained blue with DAPI. Magnification: ×200. Scale bar = 50 μm. Significance: *P < 0.05, **P < 0.01 vs. the control group. ^#P < 0.05 vs. the CAWS group