Figure 3. Aberrant epithelial necroptosis contributes to heightened colitis in Tsc1^IEC-KO mice.

Tsc1^fl/fl and Tsc1^IEC-KO mice were treated with 2.5% DSS for 5 days and then with regular water. (A and B) Body weight (A) and survival (B) of the treated mice (n = 16–18) were documented daily. (C) Leakage of FITC-dextran from the GI tract to the bloodstream was measured in DSS-treated mice for 3 days (n = 6). (D) qPCR analysis of proinflammatory gene expression (normalized to β-actin) in colons treated with DSS for 5 days (n = 6). (E) Immunoblots of colonic epithelial lysates from mice treated with DSS for 5 days (n = 3). p-RIPK1 (S166), p-RIPK3 (S232), and p-MLKL (S345) antibodies were used. Numbers under Western blot bands represent relative quantifications over actin. (F) IF staining on colon sections from various genotypes of mice treated with DSS for 5 days (n = 6). Double-positive cells were counted over 5 high-power fields (×400) per sample (n = 30). Scale bars: 20 μm. (G) Body weight of mice treated with DSS for 5 days (n = 7–8). Data were pooled from 3 independent experiments (A, B, and G) or are representative of 3 independent experiments (C–F) and are shown as mean ± SEM. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001, by log-rank test (B), unpaired Student’s t test (C, D, and G), or 1-way ANOVA (F).