Donor Dendritic Cells Regulate Immune Reconstitution after Allogeneic Bone Marrow Transplantation.

Background: Infection, graft versus host disease (GVHD) and leukemia relapse following allogeneic bone marrow transplantation (BMT) represent complications due to impaired or inappropriate immune reconstitution. Clinical data indicate that donor type 2 dendritic cells (DC) may inhibit graft-versus-leukemia effects and chronic GVHD following allogeneic BMT from HLA matched siblings (Waller et al. Blood 2001 97:2948) and more rapid recovery of lymphocytes, _{γ δ} - T-cells, and dendritic cells is associated with improved post-transplant relapse-free survival. We developed a murine BMT model to test the hypothesis that donor DC regulate immune reconstitution following BMT. B10BR [H-2k^k] recipients transplanted with CD11b-depleted donor BM cells and purified Thy 1.1⁺ congenic spleen T-cells from C57BL/6 [H-2k^b] donors had markedly enhanced expansion of donor Thy 1.1⁺ T-cells and leukemia-free survival (50%) compared to recipients of donor T-cells plus un-manipulated BM (0% survival; Li and Waller BBMT 2004 10:540). In this report we investigated whether FACS purified donor BM DC subsets would interact with donor T-cells to regulate post-transplant immune reconstitution after MHC mis-matched BMT.

Methods: B10.BR mice were lethally irradiated then transplanted with 5,000 FACS sorted Lin⁻ sca-1⁺ c-kit⁺ C57BL/6 hematopoietic progenitor cells, 300,000 Thy 1.1⁺ C57BL/6 T-cells and 50,000 of either CD11b⁺ or CD11b⁻ BM DC subsets (lin⁻ CD11c⁺). Control mice received donor T-cells and 5 x 10⁶ un-manipulated BM containing an equivalent number of donor HPC and DC. Engraftment and chimerism were assessed by FACS analysis of peripheral blood and BM. RT PCR measured levels of transcription factors in the BM at the time of hematopoietic engraftment.

Results: Equivalent survival (60%) was seen among recipients of FACS purified donor T-cells combined with FACS purified HPC alone, HPC plus CD11b⁺ and HPC plus CD11b⁻ DC. Recipients of HPC, T-cells, plus CD11b⁺ donor DC showed a large expansion of donor-spleen derived Thy 1.1⁺ regulatory T-cells (Treg, CD3⁺ CD4⁺ CD25⁺ CD69⁻) at day +10, mixed chimerism, and markedly suppressed levels of donor Thy 1.1⁺ T-cell engraftment at day +60 post-transplant compared to recipients of HPC plus T-cells. Recipients of HPC, T-cells and CD11b⁻ DC had few donor Treg at day +10, more donor CD11b⁻ blood DC, and full donor chimerism, with subsequent expansion of donor-derived T-cells over the next 2 months without developing significant GVHD. T-cell receptor (TCR) phenotyping demonstrated higher numbers of donor Thy 1.1⁺_{γ δ} - T-cells at day +30 and +60 among recipients of CD11b⁻ DC. RT PCR of mRNA from day +10 BM shows enhanced levels of Th1/Tc1 associated transcription factors JAK1, JAK2, Stat1, Stat 4 and TYK2 among transplant recipients that received FACS purified HPC, CD11b⁻ DC and T-cells compared to recipients of HPC, CD11b⁺ DC and T-cells.

Conclusion: Using highly purified fractions of donor HPC, DC, and T-cells, we demonstrate a novel and significant immune-inhibitory effect of donor CD11b⁺ DC on post-transplant immune reconstitution. In contrast, enhanced immune reconstitution was observed among recipients of FACS purified CD11b⁻ donor DC compared to grafts containing only HPC and FACS sorted donor T-cells. These data are consistent with a model of indirect allo-antigen presentation by donor DC to donor T-cells in the first days post-transplant leading to stable DC-polarization of donor T-cells that affects long-term post-transplant immune reconstitution and leukemia-free survival.