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. 2019 Sep 20;10(10):700. doi: 10.1038/s41419-019-1938-x

Fig. 6. ROS drives NLRP3-inflammasome activation in xiap−/− macrophages.

Fig. 6

a Cell ROX staining of WT and xiap−/− macrophages in the presence of TNC-TNF and ROS scavengers NAC (3 mM) and BHA (50 μM). Data are displayed as fold change over WT. b BMDMs from WT and xiap−/− were treated overnight with TNC-TNF and/or BHA. Cell death was assessed by PI uptake on flow cytometry. c Cell lysates and supernatant of macrophages treated with TNC-TNF and/or BHA (50 μM) were immunoblotted and probed for gasdermin D, NLRP3, caspase-11, caspase-1 and IL-1β. Western blots are a representative of at least three independent experiments. d, e BMDMs were treated overnight with TNC-TNF in combination with (d) BHA (50 μM) or (e) caspase-1 inhibitor (VX-765; 50 μM). IL-1β, IL-18 and IL-6 were measured in the supernatant by multiplex. f BMDMs from WT, xiap−/− and xiap−/−nlrp3−/− were treated with TNC-TNF and assayed for IL-1β, IL-18 and IL-6 by multiplex. No IL-18 was detected. g Cell lysates and supernatant of macrophages treated with TNC-TNF and/or Nigericin (10 μM) or ATP (3 mM) were immunoblotted and probed for caspase-1 and gasdermin D. Western blots are a representative of at least three independent experiments. Data shown are mean ± SEM, including n = 3 biological replicates. The experiment was repeated three times independently. Statistical significance was calculated using two-way ANOVA with *p < 0.05, **p < 0.01, ***p < 0.001 and ****p < 0.0001.