IL-38 Alleviates Inflammation in Sepsis in Mice by Inhibiting Macrophage Apoptosis and Activation of the NLRP3 Inflammasome

Figure 5

IL-38 downregulation impacts polarization and inflammasome activation and apoptosis in LPS-treated macrophages. (a) Representative western blot image (left panels) and quantification (right panels) of indicated proteins in macrophages treated as indicated. β-Actin protein levels were used for normalization. (b) Representative flow cytometry plots of macrophages treated with PBS or LPS (1 μg/mL; 24 h) alone or with anti-IL-38 siRNA in order to quantify populations with the M1 phenotype (F4/80⁺ CD11b⁺) or M2 phenotype (F4/80⁺ CD206⁺). (c) Representative western blot image (upper panels) and quantification (lower panels) of indicated proteins in macrophages treated as indicated. β-Actin protein levels were used for normalization. (d) Representative immunofluorescence images of NLRP3 (green) of macrophages treated as indicated. Nuclei were stained with DAPI (blue). Magnification, ×600. . (e, f) Representative flow cytometry plots of macrophages treated with PBS or LPS (1 μg/mL; 24 h) alone or with recombinant IL-38 (200 ng/mL; 24 h) in order to evaluate apoptosis (left panel). Percentages of apoptotic (Annexin V+) macrophages treated as indicated (right panel). Representative western blot image (left panels) and quantification (right panels) of the indicated proteins in macrophages stimulated with LPS (1 μg/mL; 24 h). β-Actin protein was used for normalization. , vs. the Ctr siRNA group. ^# and ^##, vs. the LPS+Ctr siRNA group. Results reflect three independent experiments.

(a)

(b)

(c)

(d)

(e)

(f)