RelA haploinsufficiency is associated with impaired NF-κB signaling and increased sensitivity to TNF. (A) Cell death, determined by annexin-V and/or fixable viability-dye staining, after fibroblasts from three controls (C) and patients 1 and 2 (Pts) were treated with TNF for 24 h. (B) Percentage of cleaved caspase8⁺ cells among the annexin-V⁺ population after fibroblasts from three controls and patients 1 and 2 were treated with TNF for 24 h. (C) Relative NF-κB luciferase activity in fibroblasts from three controls and patients 1 and 2 after treatment with TNF. (D) IL-6 secretion from fibroblasts and (E) PBMCs from three controls and patients 1 and 2 in response to stimulation with Pam3CSK4 (TLR1/2 agonist), LPS (TLR4 agonist), flagellin (FLA, TLR5 agonist), CL075 (TLR7/8 agonist), TNF, or IL-1β. (F) IL-10 secretion after LPS stimulation of PBMCs from patients 1 and 2 and five controls. (G) Cell death in TNF-treated PBMCs from three controls and Patients 1 and 2. All data are pooled from 3 independent experiments. Columns and bars represent means ± SEM, n.s., not significant; *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001; Student’s t test.