Donor DC reconstitution after BMT and their effect on GVHD. B6D2F1 mice received transplants of WT bone marrow with or without purified WT T cells, and splenocytes were harvested at weekly intervals. (A) DCs were enriched by density-gradient centrifugation and stained with class II-FITC, H2D^d-PE, CD11c-allophycocyanin, and H2D^d-neg CD11c^hi DCs were gated and enumerated. Results represent mean ± SEM, n = 3/group at each time point. Recipients of TCD BM (□) and BM + T (■) cells are represented. (B) B6D2F1 mice received transplants of c-fms/EGFP “MacGreen” bone marrow supplemented with or without purified WT T cells, and GFP⁺ Langerhans cells in ear pinnae were visualized by confocal microscopy (Leica Microsystems, Watzlar, Germany) at times indicated. Naive refers to an untreated MacGreen mouse. (C) B6D2F1 recipients were given transplants of CD11c-DTR Tg TCD BM with or without WT T cells and treated with DT or saline (arrows) as described in “Materials and methods.” To examine depletion efficiency and subsequent DC reconstitution, 24 hours and 10 days after the final DT injections, spleens were harvested, density-gradient–enriched, and stained with PDCA-1–PE, 7AAD, and CD11c-allophycocyanin. Shown are TCD BM recipients. (D) GVHD clinical scores were determined weekly as described in “Materials and methods.” Arrows indicate time of DT or saline injections. Data represent mean ± SEM of individual animals. *P < .05, **P < .01; allo + DT versus allo + saline recipients (n = 11-15/group).