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. 2022 Oct 12;23(20):12154. doi: 10.3390/ijms232012154

Figure 3.

Figure 3

A secondary stimulus is required for the secretion of NLRP3-dependent IL-1β in the human GEN2.2 pDC cell line. GEN2.2 cells were pretreated with 1 μM CpG-A, 1 μM CpG-B or 5 μg/mL IMQ for 3 h, then 20 μM nigericin was added to the cells in a time-dependent manner (A). In parallel experiments, cells were incubated in the presence of 1 μM MCC950 (specific NLRP3 inhibitor) for the last 30 min of TLR stimulation prior to nigericin exposure (A). In similar experiments, GEN2.2 cells were primed with live E. coli (MOI 10), Bacillus subtilis (MOI 10), Lactobacillus rhamnosus (MOI 10) and Candida albicans (MOI 0.01), then nigericin was added to the cells in a time-dependent manner (B). Similarly, in a parallel experiment, cells were incubated with MCC950 for the last 30 min of the microbial stimulation prior to nigericin treatment (B). IL-1 β secretion was measured by ELISA (A,B). Data are represented as mean ± SD of 6 individual experiments and were analyzed using one-way ANOVA followed by Bonferroni’s post-hoc test. ** p < 0.01, *** p < 0.0001, **** p < 0.0001 vs. control (ctrl), # p < 0.05, ## p < 0.01, ### p < 0.0001, #### p < 0.0001. ND: not determined, IMQ: imiquimod, NIG: nigericin, EC: Escherichia coli, BS: Bacillus subtilis, LR: Lactobacillus rhamnosus, CA: Candida albicans.