Immune responses are suppressed in the presence of GVHD. (A) A total of 10⁷ B6.WT TCD BM (non-GVHD) + 0.2 × 10⁶ CD3⁺ B6.WT T cells (GVHD) were transferred into lethally irradiated (900 cGy, day 0) BALB/c recipients on day 0. GVHD clinical scores are shown. (B) TEa Tg T cells were sorted, CFSE labeled, and injected at days 7, 14, and 35 after transplantation. CFSE dilution in spleen was analyzed 3 days later, and proliferation indices are shown. Data are representative of 3 replicate experiments, with minimum n = 4/group/time point. *P = .05 to .01. ***P < .001. (C) Representative histograms of CFSE dilution in the TEa Tg T-cell population (day 10, after day 7 injection). Syngeneic control histograms are included (from BALB/c → BALB/c transplants). (D) Percentage of dead TEa Tg T cells on day 10 was measured with 7-aminoactinomycin D. Data are representative of 4 replicate experiments: n = 16 for the non-GVHD arm; n = 19 for the GVHD arm. (E) Naive and memory CD3⁺ donor T cells (sort purified on the basis of CD62L and Thy1 expression) were transferred on day 0. TEa T cells were injected on day 7 to measure antigen presentation. Data are representative of 2 replicate experiments: n = 7 for the TCD group; n = 9 for each naive and memory T-cell arm. ***P < .001. ns indicates not significant. (F) Grafts were performed as described, and luciferase-expressing TEa Tg T cells were transferred on day 7 after transplantation. Five days later, animals were injected with luciferin, and images were acquired. (G) Bioluminescent data are combined from 2 replicate experiments: n = 7 to 9/group. *P = .05 to .01. (H-I) 10⁷ B6.CD11c.OVA BM (non-GVHD) + 0.2 × 10⁶ CD3⁺ B6.WT T cells (GVHD) were transferred into BALB/c recipients. On day 7 and 34 after transplantation, CFSE-labeled CD4⁺ OT-II T cells were transferred and proliferation was assessed 3 days later: day 7, P = .0286; n = 4/group; day 34, P = .0079; n = 5/group. Representative CFSE dilution histograms are shown in panel H. *P = .05 to .01. **P = .01 to .001. (J-K) B6.pfp^−/− grafts (10⁷ TCD B6.pfp^−/− BM ± 0.2 × 10⁶ B6.pfp^−/−CD3⁺ T cells) were transferred into BALB/c recipients. On day 7 after transplantation, third-party DBA/1 CD4⁺ T cells were CFSE labeled, adoptively transferred, and analyzed 3 days later: n = 4/group. Representative CFSE dilution histograms are shown in panel J.