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Chemokines and chemokine receptors play critical roles in directing the migration of alloreactive donor T cells into graft-vs-host disease (GVHD) target organs. However, blockade of GVHD by antagonist Ab against chemokine receptors remains an elusive goal. Using a mouse model of human GVHD, we demonstrate that in vivo administration of anti-CXCR3 Ab for 21 days (long-term), but not for 7 days (short-term), inhibits alloreactive CD8+ T cell-mediated GVHD. During a graft-vs-host reaction, infused donor CD8+ T cells generate two subsets of potent inducers of GVHD: CXCR3+CD8+ and CXCR3−CD8+ T cells. Compared with CXCR3+CD8+ T cells, CXCR3−CD8+ T cells produce less granzyme B, Fas ligand, IFN-γ, and TNF-α. Interestingly, stimulation with either dendritic cells or IL-2 induces a dynamic conversion between CXCR3+CD8+ and CXCR3−CD8+ T cells. Short-term anti-CXCR3 Ab treatment inhibits only CXCR3+CD8+ T cell-mediated GVHD, but not the disease induced by CXCR3−CD8+ T cells. Prolonged in vivo administration of anti-CXCR3 Ab significantly reduces the infiltration of alloreactive CD8+ T cells into GVHD target organs and inhibits GVHD mediated by either CXCR3+CD8+ or CXCR3−CD8+ T cells. Thus, we have established a novel and effective approach with the potential to give rise to new clinical methods for preventing and treating GVHD after allogeneic hematopoietic stem cell transplantation.

Acute and chronic graft-versus-host disease (GVHD) remain the major complications limiting the efficacy of allogeneic hemopoietic stem cell transplantation. Chronic GVHD can evolve from acute GVHD, or in some cases may overlap with acute GVHD, but how acute GVHD evolves to chronic GVHD is unknown. In this study, in a classical CD8+ T cell-dependent mouse model, we found that pathogenic donor CD4+ T cells developed from engrafted hemopoietic stem cells (HSCs) in C57BL/6SJL(B6/SJL, H-2b) mice suffering from acute GVHD after receiving donor CD8+ T cells and HSCs from C3H.SW mice (H-2b). These CD4+ T cells were activated, infiltrated into GVHD target tissues, and produced high levels of IFN-γ. These in vivo-generated CD4+ T cells caused lesions characteristic of chronic GVHD when adoptively transferred into secondary allogeneic recipients and also caused GVHD when administered into autologous C3H.SW recipients. The in vivo generation of pathogenic CD4+ T cells from engrafted donor HSCs was thymopoiesis dependent. Keratinocyte growth factor treatment improved the reconstitution of recipient thymic dendritic cells in CD8+ T cell-repleted allogeneic hemopoietic stem cell transplantation and prevented the development of pathogenic donor CD4+ T cells. These results suggest that de novo-generated donor CD4+ T cells, arising during acute graft-versus-host reactions, are key contributors to the evolution from acute to chronic GVHD. Preventing or limiting thymic damage may directly ameliorate chronic GVHD.

Host APCs are required for initiating T cell-dependent acute graft-vs-host disease (GVHD), but the role of APCs in the effector phase of acute GVHD is not known. To measure the effect of tissue-resident APCs on the local development of acute GVHD, we selectively depleted host macrophages and DCs from the livers and spleens, but not from the skin, peripheral lymph nodes (PLN), or mesenteric lymph nodes (MLN), of C57BL/6 (B6) mice by i.v. administration of liposomal clodronate before allogeneic bone marrow transplantation. Depletion of host hepatic and splenic macrophages and DCs significantly inhibited the proliferation of donor C3H.SW CD8+ T cells in the spleen, but not in the PLN or MLN, of B6 mice. Such organ-selective depletion of host tissue APCs also markedly reduced the trafficking of allogeneic CD8+ T cells into the livers and spleens, but not PLN and MLN, of B6 recipients compared with that of the control mice. Acute hepatic, but not cutaneous, GVHD was inhibited as well, resulting in improved survival of liposomal clodronate-treated B6 recipients. When C3H.SW CD8+ T cells were activated in normal B6 recipients, recovered, and adoptively transferred into secondary B6 recipients, activated donor CD8+ T cells rapidly migrated into the livers and spleens of control B6 recipients but were markedly decreased in B6 mice that were depleted of hepatic and splenic macrophages and DCs. Thus, tissue-resident APCs control the local recruitment of allo-reactive donor T cells and the subsequent development of acute GVHD.