Fig. 6: 3-HKA inhibits IFN-γ-mediated STAT1/NFκB activation.

a Representative immunoblot analysis of STAT1 and the NF-κΒ pathway-associated proteins p65 and p50 from human monocyte-derived DCs treated or untreated with 1 µM 3-HKA with or without 50 ng/ml IFN-γ for 20 min. b Representative immunoblot analysis of pSTAT1 from human monocyte-derived DCs treated or untreated with titrated amounts of 3-HKA with or without 50 ng/ml IFN-γ at different time points. c, d, e Densitometric analysis of triplicate immunoblots as reported in (a) and (b). Data of n = 3 biologically independent replicates are plotted as mean relative expression ± SD. Significance levels are reported as p < 0.05 (*), p < 0.01 (**), and p < 0.0001 (****) (two-way ANOVA followed by Tukey’s multiple comparison test). f Representative immunoblot analysis of pIκΒα from human monocyte-derived DCs treated or untreated with 1 µM 3-HKA with or without 50 ng/ml IFN-γ for 20 min. g Densitometric analysis of triplicate Western blots as reported in (g). Data of n = 3 biologically independent replicates are plotted as mean relative expression ± SD. Significance levels are reported as p < 0.05 (*), p < 0.01 (**), and p < 0.001 (***) (two-way ANOVA followed by Tukey’s multiple comparison test). h, i IPA-predicted quantitative decrease of proteins associated with the IFN-γ-mediated signal transduction in mouse DCs treated with 3-HKA and IFN-γ as compared with IFN-γ treatment alone (generated from data reported in Supplementary Data 2). j Scheme of the delayed hypersensitivity skin test assay. k In vivo suppression activity of HY-pulsed CD8⁻CD11c⁺ DCs (cDC) in combination with a minority fraction (5%) of the same DCs with no conditioning or conditioned in vitro with 1 µM 3-HKA for 24 h. Analysis of the delayed hypersensitivity skin reactivity of recipient mice to the eliciting peptide at 15 days is presented as change in footpad weight (experimental versus control footpads). Data are reported as average footpads weight ± SD of n = 6 biologically independent replicates. Data were analyzed by paired T test (Wilcoxon test), ***p < 0.001. Source data for (a–g) and (k) are provided as Source data file.