Figure 6. LTα₃ can induce both CASP8-dependent apoptosis and RIPK3-dependent necroptosis in A20/Abin-1^T-ΔIEC Tnf^–/– enteroids.

(A and C) Quantitative luminescent cell viability assay of enteroids with the indicated genotypes treated with vehicle, 4-OHT, Nec1s, or emricasan for 24 hours as indicated, and then treated with the indicated stimuli for 24 hours (mean ± SEM; 50 μM Nec1s, emricasan; 50 ng/mL TNF). (B and D) Immunoblot analyses of enteroid cultures with the indicated genotypes treated with 4-OHT for 22.5 hours, and then vehicle, Nec1s, or emricasan for 1.5 hours, followed by 20 ng/mL recombinant human LTα₃ (hLTα₃) as indicated. Lysates were immunoblotted with the antibodies indicated on the right. Solid arrows indicate full-length protein; open arrows indicate cleaved protein. For panel A, significance was assessed using 2-way ANOVA with Dunnett’s multiple-comparison test relative to LTα₃-Fc CM plus Nec1s. For panel C, significance was assessed by 2-way ANOVA with Bonferroni’s multiple-comparison test comparing between genotypes for each stimulation condition. Only significant differences are shown. ****P < 0.0001. Data represent at least 2 independent experiments.