Etiology of anemia in a murine model of myelodysplastic syndrome

Abstract

Abstract MDS are clonal stem cell disorders characterized by peripheral cytopenias, particularly macrocytic anemia, which is often refractory to erythropoiesis-stimulating agents, necessitating chronic transfusions that contribute to poor quality of life and increased mortality. To better understand the etiology of anemia in MDS we investigated the erythroid lineage in the Vav-driven NUP98/HoxD13 (NHD13) murine model, which faithfully recapitulates many aspects of MDS including the development over time of macrocytic anemia and progression to acute leukemia. The steady-state production of red blood cells depends on expansion of erythroid progenitors, as well as progressive doubling of terminally maturing erythroblasts. Analysis of anemic NHD13 mice revealed not only decreased numbers of erythroid progenitors but also, surprisingly, a failure of terminal erythroblast expansion. As MDS is a clonal disorder, we generated chimeric NHD13 mice by transplanting wildtype (WT) or NHD13 marrow along with WT UBC-GFP+ marrow into WT recipient mice. While non-anemic WT:GFP+ WT mice maintained stable ratios of terminal erythroblasts, NHD13:GFP+ WT (NHD13) chimeric mice evidenced a failure of clonal (NHD13) terminal erythropoiesis, consistent with an intrinsic defect in terminal NHD13 erythropoiesis. However, this defect was compensated by the WT cells resulting in the majority of circulating red blood cells being WT (GFP+). Interestingly, the development of macrocytic anemia in chimeric NHD13 mice was associated with terminal failure not only of clonal erythroblasts but also of WT erythroblasts. These findings are consistent with a microenvironmental impact on WT erythroid precursors in the setting of progressive disease. Since erythroblasts terminally mature within erythroblastic islands in association with myeloid cells, we analyzed the numbers of myeloid cells in the bone marrow of NHD13 chimeric mice and found a 50% reduction of macrophages, as well as a four-fold expansion of monocytes, almost all of which were derived from the NHD13 clone. We developed an imaging flow cytometric assay to characterize erythroblast-myeloid cell interactions in chimeric NHD13 mice and found reduced macrophage-erythroblast associations and increased monocyte-erythroblast associations. Compared to WT marrow monocytes, monocytes from NHD13 mice expressed increased levels of multiple inflammation-associated genes including IFNg and IL1b. Consistent with the upregulation of IL1bR in terminal erythroblasts, IL1b treatment inhibited the terminal in vitro maturation of human CD34-derived, as well as primary murine, erythroblasts. Taken together, our studies in Vav-driven constitutive and chimeric NHD13 mice indicate that an NHD13-associated cell intrinsic defect in terminal erythropoiesis is initially compensated by WT erythropoiesis, which subsequently fails with progression to anemia. The erythroid precursors in the marrow of anemic NHD13 chimeric mice have increased interactions with (clonal) monocytes, which are more inflammatory than WT monocytes and likely inhibit WT terminal erythropoiesis, mediated in part by IL1b. We conclude that the etiology of anemia in the NHD13 model of MDS is complex having both cell intrinsic and cell extrinsic components.