Fig. 4. Gut-aGVHD induced by IFN-γ^−/− donor CD8⁺ T cells is associated with expansion of Tc17 and Tc22 cells.

Lethally irradiated WT BALB/c recipients were engrafted with splenocytes containing 1.5 × 10⁶ T cells combined with TCD-BM (2.5 × 10⁶) from WT or IFN-γ^−/− C57BL/6 donors. Additional controls include IFN-γ^−/− C57BL/6 grafts in syngeneic WT C57BL/6 recipients and MHC I-matched H-2K^bMHC-IA⁻-IE⁻BALB/c recipients. Allogeneic recipients were also treated with depleting anti-CD4 mAb (500 µg/mouse) immediately after HCT (a–c). a Mean ± SEM of %Original bodyweight at each time point and recessive curves of %mice without diarrhea and %survival among total mice are shown. n = 8 (syngeneic & MHC I-matched recipients), 10 (IFN-γ^−/− CD8⁺ & WT CD8⁺). b Seven and 14 days after HCT, histopathology of colon was evaluated. Representative micrographic photos and %mice with indicated colitis histopathology scores are shown; n = 4 biologically independent mice per group. c At day 7 after HCT, MLN of recipients were analyzed for donor-type IL-17A⁺IL-22⁻, IL-17A⁺IL-22⁺ or IL-17A⁻IL-22⁺ CD8⁺ T-cell subsets. Means ± SE of percentages are shown, n = 10. d–f Lethally irradiated WT BALB/c recipients were engrafted with sorted CD8⁺ T (1.5 × 10⁶) from IFNγ^−/− or IFNγ^−/−/RORγt^−/− C57BL/6 donors combined with TCD-BM (2.5 × 10⁶) from WT C57BL/6 donors. d Recessive curves of %mice without diarrhea and %survival among total mice are shown. n = 8 from two replicate experiments. e, f On day 7 after HCT, MLN cells were analyzed for %IL17A⁻IL-22⁺ CD8⁺ T cells (e) and %AHR⁺RORγt⁻ or AHR⁻RORγt⁺ among IL17A⁻IL-22⁺CD8⁺ T cells (f) means ± SEM are shown, n = 4. All results combined from two replicates. Each dot represents one mouse. Log-rank test was performed with two-tailed p value for survival comparisons. Nonlinear regression (curve fit) was used and a two-tailed p value was calculated for bodyweight and diarrhea comparisons. Unpaired two-tailed Student’ t test was used to compare means. a ****p < 0.0001; c **p = 0.0063, ***p = 0.0006; d ****p < 0.0001. e **p = 0.0010 (IFNγ^−/−), **p = 0.0059 (IFNγ^−/−/RORγt^−/−). T-cell-depleted bone marrow (TCD-BM), SPL (spleen), MLN (mesenteric lymph node).