Hemolysis and the Hematopoietic Niche

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ABSTRACT Sickle cell disease (SCD) is a major healthcare burden with limited treatment options. Currently available treatments are limited to transfusions, hydroxyurea and L-glutamine. With increasing interest in hematopoietic stem cell (HSC) transplantation and gene therapy as treatment options for SCD, understanding how hemolytic stress alters the bone marrow (BM) microenvironment is critical for the development of appropriate therapeutic strategies. In Aim 1, we will investigate the effects of hemolysis on the BM niches including mesenchymal stem cells (MSCs), responsible for maintaining HSCs, and erythroblastic islands (EBIs), the niche for erythropoiesis. In Aim 2, we will define the mechanisms for the effects of hemolysis on erythroid progenitors/precursors. We further propose to determine whether therapy with transfusion, heme scavenger hemopexin or the combination can improve the function of MSCs and EBI macrophages. Our overall hypothesis is that free heme leads to dysfunction of the BM hematopoietic niche and hematopoietic stem/progenitor cells, which can be alleviated in part by hemopexin and/or transfusion therapy. First, we will investigate the effects of hemolysis on the BM MSC ability to regulate hematopoiesis. We will evaluate numerous MSC stem cell properties and investigate how alterations may in turn affect HSC activation state, accumulation of ROS and DNA damage. Next, we will define the mechanisms for the impaired erythropoietic activity due to hemolysis, including mechanisms for defective EBIs formation, CFU-E colony formation and enucleation. These studies are enabled by the SCD mouse model, Thal mouse model, Epor-eGFP knockin mouse model, mouse models targeting BM MSCs, our recent identification of Epor+ EBI macrophages, validation of both anti-mouse Epor and anti-human EPOR antibodies as well as the methods we have developed to purify erythroid cells at distinct developmental stage and to quantify both murine and human BM terminal erythroid differentiation. These studies will provide a comprehensive mechanistic understanding of the effects of hemolysis on BM hematopoietic niches, and contribute to the development of novel therapeutics targeting the BM niche cells and ultimately improve treatment options for patients with SCD.