Mice are the primary animal of choice for modeling aGVHD. One aspect of these mouse models that may limit translation of promising therapeutic strategies to patient treatment is the use inbred mice as surrogates for genetically diverse human populations. Objective Quantify and compare tissue inflammation and injury in inbred vs. genetically diverse outbred mice using a well-characterized model of aGVHD-mediated bone marrow (BM) failure and spleen hypoplasia. Methods C57Bl6 (Bl6) CD4+ T cells (20,000 T cells/g b.w.) were injected (i.p.) into sub-lethally irradiated Bl6-H2-Ab1bm12(BM12) recipients (Bl6→BM12) or into Collaborative Cross (CC) or CD1 outbred recipients. Results Adoptive transfer of allogeneic Bl6 T cells into BM12 recipients induced marked weight loss (25%) at 20 days post T cell transfer that was associated with dramatic reductions in hematocrit as well as large and significant decreases in circulating granulocytes, platelets and erythrocytes when compared to their syngeneic controls (BM12→BM12). Total cell numbers in BM and spleen were also found to be dramatically reduced in these mice. Histopathological inspection confirmed severe hypocellularity in BM and spleen whereas no aGVHD was present in the lungs, liver, skin and colon. In contrast, adoptive transfer of Bl6 T cells into sub-lethally irradiated CC or CD1 or CC mice induced little or no weight loss at 25–30 days post T cell transfer with no significant reductions in hematocrit, circulating leukocytes, erythrocytes or platelets. BM- and spleen-associated cell numbers were not reduced when compared their CC controls. Conclusions Genetically diverse outbred mice are resistant to aGVHD-mediated BM failure and spleen hypoplasia.

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Allogeneic hematopoietic stem cell transplantation (HSCT) is a potentially life-saving treatment for refractory/relapsing hematological malignancies, blood disorders or autoimmune diseases. However, approximately 40–50% of patients undergoing allogeneic HSCT will develop a multi-organ, inflammatory disorder called acute graft vs. host disease (aGVHD). Experimental and clinical studies suggest that intestinal injury due to toxic, pre-transplant conditioning protocols (e.g. lethal irradiation and/or chemotherapy) may play a major role in the development of aGVHD. However, recent studies from our laboratory as well as others, suggest that this may not be the case. Objective We wished to quantify and compare the onset and severity of aGVHD induced by the adoptive transfer of allogeneic T cells into untreated lymphopenic mice. Methods Four million allogeneic or syngeneic CD4+CD62L+CD25− T cells were transferred (i.p.) into NK cell-depleted RAG1−/− mice or RAG2−/−IL2rγ−/− double knock-out (DKO) mice and assessed daily for signs of aGVHD. Results We found that adoptive transfer of allogeneic but not syngeneic T cells into NK cell-depleted RAG1−/− or DKO mice induced many of the clinical and histological features of aGVHD including weight loss, inflammatory cytokine production and tissue inflammation. In addition, adoptive transfer of allogeneic T cells into each recipient induced severe anemia as well as dramatic reductions in bone marrow and spleen cellularity. Conclusions We conclude that naïve allogeneic CD4+T cells are both necessary and sufficient to induce aGVHD in lymphopenic recipients in the absence of toxic, pre-transfer conditioning.

Acute graft vs. host disease (aGVHD) is a major complication following hematopoietic stem cell transplantation. Preclinical and clinical studies suggest aGVHD may be potentiated by gut damage and translocation of intestinal microbiota following toxic, pre-transplant conditioning protocols. We recently reported that aGVHD-induced bone marrow (BM) failure and splenic hypoplasia develops in the absence of gut damage suggesting that intestinal bacteria may not be required for disease pathogenesis. Objective Determine whether prophylactic gut decontamination with broad spectrum antibiotics (Abx) affects the onset and/or severity of aGVHD-induced BM and spleen damage. Methods Syngeneic (Bl6) or allogeneic (Balb/c) CD4+CD25−T cells (5×106 cells) were injected into NK cell-depleted Bl6 RAG1−/− recipients. Prior to T cell transfer, RAG1−/− mice received water (ab libitum) containing aspartame (Asp) or an Abx cocktail containing Asp, neomycin and vancomycin for 7 days prior to and following T-cell transfer. Results Treatment of allogeneic mice with Abx reduced colonic bacterial load by more than 20-fold when compared to their Asp-treated counterparts. Abx treatment also resulted in large and significant reductions in BM- and spleen-residing T cells and myeloid cells as well as circulating erythrocytes, platelets and hematocrit when compared to Asp treated mice. These Abx-induced alterations were associated with significant increases (~4-fold) in serum IL-6 levels compared to Asp-treated mice. Conclusions Prophylactic Abx treatment exacerbates aGVHD-induced BM failure and spleen hypoplasia.