Allogeneic T cells induce cell death in intestinal organoids with autophagy gene mutations. Representative images (A), viability (B), and size (C) of small intestinal organoids from B6-background Atg16L1^f/f (f/f) and Atg16L1^ΔIEC (ΔIEC) mice cocultured for 48 hours with 1 × 10⁵ splenic T cells separately harvested from B6, B10.BR, and LP/J mice. n = 3 mice each. Arrowheads indicate dead organoids. Scale bars, 400 µm. (D) Viability of organoids from B6-background Atg4B^−/− and Atg16L1^T316A mice cocultured for 48 hours with 1 × 10⁵ splenic T cells separately harvested from B10.BR mice; n = 3 mice each. (E) Viability of small intestinal organoids from f/f and ΔIEC mice cocultured for 48 hours with FACS-sorted 1 × 10⁵ CD4⁺ or 7 × 10⁴ CD8⁺ T cells from B10.BR and LP/J mice; n = 3 mice each. Representative images (F) and number of T cells associated with organoid (G). At least 50 organoids were analyzed per group. T cells were stained with CellBrite Green (green) before coculture, and propidium iodide (PI; red) was added to the culture medium at the beginning to stain dead organoids/T cells. Scale bars, 25 µm; n = 3 mice each. Data points in B, D, and E are mean of technical replicates, and data points in C and F represent individual organoids. Bars represent mean ± standard error of the mean, and ≥2 independent experiments were performed. *P < .05, **P < .01, ***P < .001, ****P < .0001. ns, not significant; WT, wild-type.