The level of residual host CD11c⁺ DCs does not influence the DLI-mediated LH-GVH response in MHC-matched in contrast to MHC-mismatched chimeras. (A-C) MHC-matched setting. (A) DC chimerism analysis of 8-week-old B10.D2 + BALB/c-CD45.1→BALB/c-CD45.1 mixed and B10.D2→BALB/c-CD45.1 full chimeras. Representative histograms show differing levels of residual host DC chimerism in the spleen (SPL) and cutaneous lymph nodes (CLNs) of mixed versus full chimeras and near complete persistence of host-derived CD11c⁺ DCs in the skin of both groups. Data represent results of 2 independent experiments. (B-C) Serial analysis of multilineage chimerism in peripheral blood after DLI administration. The 8-week-old B10.D2 + BALB/c-CD45.1→BALB/c-CD45.1 mixed chimeras received nothing (□; n = 5), 2 × 10⁷ splenocytes from naive B10.D2 donor mice (▪; n = 5) or the same dose of DLI from B10.D2 mice presensitized to host antigens (▵; n = 5). 8-week-old of B10.D2→BALB/c-CD45.1 full chimeras received nothing (⋄; n = 5) or 2 × 10⁷ splenocytes from the same naive donor source (♦; n = 5). Changes in donor CD4⁺ T-cell (B) and granulocyte chimerism (C) were monitored by serial tail bleeding and flow cytometric analysis using fluorochrome-conjugated antibodies against Gr-1, CD4, and CD45.1. Data are presented as a mean percentage of donor chimerism ± SEM and represent one of 2 independent experiments. (D-G) MHC-mismatched setting. At 8 weeks after alloBMT MHC-mismatched B6 + BALB/c→BALB/c mixed chimeras received nothing (open symbols) or DLI (filled symbols) in the form of 2 × 10⁷ splenocytes from donor B6 mice. (D) Changes in CD4⁺ (□, ▪) and CD8⁺ (⋄, ♦) T-cell chimerism were determined in peripheral blood by flow cytometry after staining with anti–H-2^b–, anti-CD4–, and anti-CD8–specific antibodies. Data are presented as a mean percentage of donor chimerism ± SEM (n = 5 mice/group). (E) At 8 weeks after alloBMT 2 × 10⁷ CFSE-labeled splenocytes from B6.PL-Thy1^a mice were transferred to B6 + BALB/c→BALB/c mixed and B6→BALB/c full donor chimeras. Representative CFSE profiles of gated DLI-derived CD4⁺Thy1.1⁺ and CD8⁺Thy1.1⁺ T cells analyzed using anti-Thy1.1–, anti-CD4–, and anti-CD8–specific antibodies on day 5 after adoptive transfer. Gating used to delineate unproliferated CFSE^hi from fast proliferating DLI-derived T-cells is indicated. (F) Percentages of gated CFSE^hi DLI-derived CD4⁺ and CD8⁺ T cells in spleens of mixed and full chimeras on day +5 after DLI. (G) Absolute numbers of IFN-γ–producing DLI-derived T cells in the spleens of mixed and full MHC-mismatched chimeras on day +5 following DLI (after brief ex vivo stimulation). Data shown represent 1 of 3 independent experiments (mean ± SEM; n = 3-4 mice/group).