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. 2017 Dec 4;214(12):3687–3705. doi: 10.1084/jem.20170558

Figure 5.

Figure 5.

ATG16L1 inhibits necroptosis in intestinal organoids. (A and B) Representative images and quantification of TUNEL+ (A) and CC3+ (B) cells per organoid on day 5 normalized to total IEC. TNFα was added on day 4. At least 20 organoids were quantified from three mice each. (C and D) Quantification of viability (C) and representative images of organoids (D) from Atg16L1f/f and Atg16L1ΔIEC mice on day 5 ± Z-VAD-FMK (Z-VAD) and Nec-1. (E) Representative TEM images of organoids on day 3. Z-VAD was added on day 0. Stars indicate lumen side of organoid, and arrows indicate Paneth cell. (F and G) Quantification of viability (F) and representative images of organoids (G) from Atg16L1f/f and Atg16L1ΔIEC mice on day 5 ± TNFα and Nec-1. (H and I) Viability of Atg16L1ΔIEC organoids transduced with lentiviruses encoding shRNAs targeting Ripk3 (H) and Mlkl (I) or a nonspecific control, ±TNFα. (A, B, D, E, and G) Bars: 50 µm (A and B); 100 µm (D and G); 2 µm (E). Data points in A and B represent individual organoids, and data points in C, F, H, and I are mean of three technical replicates. Bars represent mean ± SEM, and at least two independent experiments were performed. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001 by one-way ANOVA and Tukey’s test in A and B and unpaired t test in C, F, H, and I.