Fig 1: Indomethacin prevents clearance of invading microbes leading to a heightened inflammatory response. (a) YAMC cells were infected with Salmonella (MOI 100) for 30 minutes, followed by addition of the appropriate compound. At the indicated time point, bacterial load was determined via flow cytometry. (b) YAMC cells were infected with Salmonella (MOI 100) for 30 minutes, followed by addition of the appropriate compound. Eighteen hours post infection, the concentration of IL-18 in the supernatant was measured and normalized to the number of cells per well. (c) Mice (n = 6/group) were administered indomethacin (10 mg/kg), streptomycin or vehicle control. The next day, mice were inoculated with 1 × 108 CFU of Salmonella Typhymurium. Twenty-four hours post infection, quantitative culture of viable bacteria in the Peyer’s patch was performed. Values and error bars represent the average and 95% confidence intervals, respectively. IM: indomethacin; Strep: streptomycin, *p < 0.05.
Fig 2: Hippo signaling suppresses the NLRP3 inflammasome activation in a YAP-dependent manner.a, b ELISA of IL-1ß, IL-18, and TNF-a in supernatants of mouse peritoneal macrophages from Yapfl/fl lyz2-Cre and Yapfl/fl mice treated with serum starvation for indicated times (a) or seeded into different confluence (b), then applied with indicated stimuli (for a, mean ± SD, two-way ANOVA with Bonferroni test, left panel:12 h and 24 h vs. 0 h in Yapfl/fl or Yapfl/fl lyz2-Cre group, ns = 0.1243, *P = 0.0267, ns > 0.9999, ns > 0.9999, **P = 0.0065, **P = 0.0055, ns > 0.9999, ns > 0.9999, ns = 0.0661, **P = 0.0069, ns > 0.9999, ns > 0.9999 in sequence; middle panel:12 h and 24 h vs. 0 h in Yapfl/fl or Yapfl/fl lyz2-Cre group, *P = 0.0448, *P = 0.0261, ns > 0.9999, ns > 0.9999, ns = 0.06, *P = 0.0113, ns > 0.9999, ns > 0.9999, ns = 0.0557, *P = 0.0127, ns > 0.9999, ns > 0.9999 in sequence; n = 3 independent experiments. For b, mean ± SD, two-way ANOVA with Bonferroni test, left panel: 12-well and 24-well vs. 6-well in Yapfl/fl or Yapfl/fl lyz2-Cre group, ns = 0.1743, *P = 0.0239, ns > 0.9999, ns = 0.5177, ns = 0.0861, *P = 0.0408, ns > 0.9999, ns > 0.9999, *P = 0.0115, **P = 0.0021, ns > 0.9999, ns > 0.9999 in sequence; middle panel: 12 h and 24 h vs. 0 h in Yapfl/fl or Yapfl/fl lyz2-Cre group, ns = 0.0969, *P = 0.0386, ns > 0.9999, ns > 0.9999, ns = 0.1, **P = 0.0038, ns > 0.9999, ns > 0.9999, *P = 0.0205, **P = 0.0032, ns > 0.9999, ns > 0.9999 in sequence; n = 3 independent experiments). c ELISA of IL-1ß, IL-18, and TNF-a in supernatants from mouse peritoneal macrophages silenced of Lats1/2 and treated with serum starvation for indicated times, then applied with indicated stimuli (mean ± SD, two-way ANOVA with Bonferroni test, si Lats1/2 24 h vs. si Ctrl 24 h, left panel: **P = 0.0031, *P = 0.0107, *P = 0.0492 in sequence; middle panel: *P = 0.0492, *P = 0.0393, *P = 0.0478 in sequence; n = 3 independent experiments). Source data are provided as a Source Data file.
Fig 3: YAP specifically promotes NLRP3 inflammasome activation.a ELISA of IL-1ß, IL-18, and TNF-a in supernatants from mouse peritoneal macrophages silenced of YAP, treated with indicated stimuli (mean ± SD, two-way ANOVA with Bonferroni test, YAP siRNA vs. Ctrl siRNA, left panel: **P = 0.0023, ***P = 0.0007, ***P = 0.0005 in sequence; middle panel: *P = 0.0222, **P = 0.0033, **P = 0.0076 in sequence; n = 3 independent experiments). b Immunoblot analysis of supernatants (SN) or cell lysates (CL) from mouse peritoneal macrophages silenced of YAP, treated with indicated stimuli. c ELISA of IL-1ß, IL-18, and TNF-a in supernatants of mouse peritoneal macrophages from Yapfl/fl lyz2-Cre or Yapfl/fl mice, then treated with indicated stimuli (mean ± SD, two-way ANOVA with Bonferroni test, Yapfl/fl lyz2-Cre vs. Yapfl/fl, left panel: ***P = 0.0003, **P = 0.0046, ***P = 0.0006 in sequence; middle panel: **P = 0.0021, **P = 0.002, **P = 0.0013 in sequence; n = 3 independent experiments). d Immunoblot analysis of supernatants (SN) or cell lysates (CL) of mouse peritoneal macrophages from Yapfl/fl lyz2-Cre or Yapfl/fl mice, then treated with indicated stimuli. e Immunoblot analysis of ASC oligomerization in cross-linked cytosolic pellets of mouse peritoneal macrophages from Yapfl/fl lyz2-Cre or Yapfl/fl mice, primed with LPS, and followed by stimulation with nigericin. f Representative images of ASC specks in LPS primed Yapfl/fl lyz2-Cre or Yapfl/fl peritoneal macrophages treated with indicated stimuli. ASC, green; nuclei, blue. White arrows indicate ASC specks. Scale bars, 10 µm (left). The percentage of cells containing an ASC speck was quantified (right). At least 100 peritoneal macrophages from each genotype were analyzed (mean ± SD, two-way ANOVA with Bonferroni test, Yapfl/fl lyz2-Cre vs. Yapfl/fl, right panel: **P = 0.0011, ***P < 0.0001 in sequence; n = 3 independent experiments). Similar results were obtained from three independent experiments. Source data are provided as a Source Data file.
Fig 4: ß-TrCP1 inhibits NLRP3 inflammasome activation.a ELISA of IL-1ß, IL-18, and TNF-a in supernatants from mouse peritoneal macrophages silenced of ß-TrCP1, treated with indicated stimuli (mean ± SD, two-way ANOVA with Bonferroni test, ß-TrCP1 siRNA vs. Ctrl siRNA, left panel: **P = 0.0024, ***P = 0.0006, ***P = 0.0006 in sequence; middle panel: *P = 0.0169, **P = 0.0011, **P = 0.0081 in sequence; n = 3 independent experiments). b Immunoblot analysis of extracts from mouse peritoneal macrophages silenced of ß-TrCP1, then stimulated for indicated times with LPS. c ELISA of IL-1ß, IL-18, and TNF-a in supernatants from Yapfl/fl lyz2-Cre or Yapfl/fl peritoneal macrophages silenced of ß-TrCP1, treated with indicated stimuli (mean ± SD, two-way ANOVA with Bonferroni test, Yapfl/fl lyz2-Cre + ß-TrCP1 siRNA vs. Yapfl/fl lyz2-Cre + Ctrl siRNA, left panel: **P = 0.0023, **P = 0.0018, **P = 0.0047 in sequence; middle panel: **P = 0.0084, **P = 0.0066, *P = 0.0498 in sequence; n = 3 independent experiments). d Immunoblot analysis of lysates from Yapfl/fl lyz2-Cre or Yapfl/fl mouse peritoneal macrophages silenced of ß-TrCP1, treated with indicated stimuli. Similar results were obtained from three independent experiments. Source data are provided as a Source Data file.
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