Figure 10. Degradation of STAT3 in donor T cells prevents acute GVHD in a PD-1–dependent manner.

(A) Splenic T cells from WT or PD-1^–/– C57BL/6 mice were treated with or without 40 μM SD-36. T cell expression of STAT3 was measured by immunoblotting after 24 hours. Representative immunoblotting patterns and means ± SEM of expression levels are shown. (B) Irradiated BALB/c recipients were engrafted with TCD-BM (5 × 10⁶) from WT C57BL/6 donors alone or with purified T cells (1 × 10⁶) from spleens of WT or PD-1^–/– C57BL/6 donors after culture in medium containing 40 μM SD-36 for 24 hours in vitro. Recipients were treated with SD-36 (50 mg/kg, i.v.) or vehicle on days 0 and 3 after HCT. Plots of percentages of original body weight and clinical GVHD score and percentage survival are shown. n = 3 (TCD-BM); n = 5 (WT T cells+vehicle); n = 6 (WT T cells+SD-36); n = 6 (PD-1^–/– T cells+SD36) combined from 2 replicate experiments. (C–H) On day 6 after HCT, lymphocytes from the liver and gut were analyzed with flow cytometry. n = 4–5 combined from 2 replicate experiments. (C) Yield of CD4⁺ donor T cells are shown. (D and E) Percentage of GM-CSF⁺IFN-γ⁺ and TNF-α⁺IFN-γ⁺ among CD4⁺ T cells. (F) Yield of CD8⁺ donor T cells. (G) Percentages of GM-CSF⁺IFN-γ⁺ among CD8⁺ T cells are shown. n = 5 combined from 2 replicate experiments. (H) MFI of GzmB of CD8⁺ T cells is shown. n = 4–5 representing means ± SEM. P values were calculated by 2-way ANOVA (A) or unpaired 2-tailed Student t tests (C–H). NS, P ≥ 0.05; *P < 0.05; **P < 0.01; ****P < 0.0001.