Figure 3.

TNFRSF receptor costimulation activated the canonical NF‐κB pathway in Tregs. (A, B) Tregs, pre‐activated with anti‐CD3/CD28 mAbs for 24 h, were restimulated with TNFRSF agonists alone. The EMSA (A) was performed at 0, 0.5, 1, and 4 h of restimulation. The supershift (B) was performed at 0.5 h of restimulation to investigate the subset composition of the NF‐κB containing complex. Arrowheads indicate the position of the NF‐κB containing complex and arrows indicate the positions of the supershifting complexes bound to mAb specific to RelA and c‐Rel. A representative of three experiments is shown. Proliferation (C, D) and survival (E) of Rela KO and control Tregs, stimulated and analyzed as in Fig. 1. One representative proliferation profile (C) and increased proliferation and FC living cell numbers relative to the control culture (D, E) from the pool of two independent experiments with two mice per experiment. Each circle represents a mouse (two‐way ANOVA, *p < 0.05, ***p < 0.001.) and the horizontal bar represents the mean. In (C), around 20 000 cells were analyzed per gated Tregs.