Fig. 6.

Genetic deficiency of SESN2 exacerbates pyroptosis and NLRP3 inflammasome hyperactivation in sepsis. DCs were isolated from the spleen at 24 h following CLP. A, Pyroptosis of DCs was analyzed by flow cytometry, and DCs were labeled with active CASP-1 (FAM-FLICA) and 7-AAD. Representative flow cytometry plots are shown on the left, and quantitative analyses of FLICA⁺ 7AAD⁺ DCs are shown on the right (n = 6). B, Representative electron micrographs (10,000 × , 50,000 ×) of morphological alterations in the cell membrane, cytoplasm, ER, and mitochondrial organelles of splenic DCs (the red arrows indicate mitochondria, the orange arrows indicate the ER, the black triangles indicate cytoplasmic vacuoles, and the black arrows indicate membrane pores). C, Immunoblot analysis of CASP-1, GSDMD, NLRP3, and ASC expression in DCs from WT and SESN2^−/− mice at 24 h after CLP. β-actin served as an internal control. D, The plasma levels of cleaved CASP-1 were measured by ELISA at 24 h after CLP (n = 5). E, Representative confocal immunofluorescence microscopy images of NLRP3 colocalized with ASC. DyLight 488 (green)-labeled ASC protein, DyLight 594 (red)-labeled NLRP3 protein, and DAPI (blue)-stained nuclei are shown. The images are representative of six independent spleen samples from the CLP group. The data are presented as the mean ± SD of at least three independent experiments. Statistical significance: ^*P < 0.05 versus the control group