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. Author manuscript; available in PMC: 2022 Oct 29.
Published in final edited form as: Nat Immunol. 2022 Apr 29;23(5):705–717. doi: 10.1038/s41590-022-01192-4

Figure 5. Procaspase-11 protein is required for assembly of the NLRP3 inflammasome in response to avirulent Gram-negative bacteria.

Figure 5.

a, Immunoblots of WT, Casp11–/– or Casp1–/– macrophage concentrated supernatants (20hr) or WCE (6hr), and cytokine concentrations and LDH release in culture supernatants (20hr) post-stimulation with L, HK or HK E. coli supplemented with E. coli RNAtot. LDH release measured by cytotoxicity assay; IL-1β and IL-6 by ELISA. b, Immunoblot of WT, Casp11–/– or Casp1–/– macrophage cross-linked fractions (16hr) post-stimulation with L E. coli. c, Immunoblots of WT, Casp11–/– or Casp1–/– macrophage concentrated supernatants (20hr) (top panels) or cross-linked fractions (16hr) (bottom panels) post-stimulation with L, HK or HK E. coli supplemented with E. coli RNAtot or poly(dA:dT). d, Immunofluorescence confocal microscopy on WT, Casp11–/– or Casp1–/– macrophages stimulated for 16hr with L E. coli RFP. Bar graph, % cells exhibiting ASC speck. Inset shows magnification of indicated area. White arrowhead points to ASC speck. Phalloidin delineates the macrophage actin cytoskeleton. Scale bar=10μm. Error bars, mean ± s.e.m. One-way ANOVA followed by multiple comparisons Sidak tests and p values indicated in bar graphs in a (IL-1β, WT and Casp1–/– : n=3, Casp11–/– : n=4; LDH, n=3) and d (WT: n=6, Casp11–/– : n=7, Casp1–/– : n=8). ns: non-significant. Bacteria:macrophage=20:1. Results represent at least 3 independent experiments.