Terminal maturation of erythroid progenitors

Blood transfusions are of utmost importance in modern medicine. Researchers all over the world try to overcome the growing supply bottleneck of healthy blood products from donors with ex vivo erythropoiesis cultures. Over the last decades, expansion, differentiation and maturation of HSPCs allowed for the generation of red blood cells in defined ex vivo culture conditions. However, ex vivo erythropoiesis lacks the final terminal maturation step from reticulocytes to biconcave erythrocytes, hindering a clinical application of cultured red blood cells so far. A detailed understanding of the underlying mechanisms and the optimal orchestration of potential triggers for this terminal reticulocyte-to-erythrocyte transition remains elusive to date. The current project aims to better define the crucial cellular interactions, soluble co-factors and flow/shear stress to prime reticulocytes for terminal transition from reticulocytes to biconcave erythrocytes ex vivo. Therefore, we extend the established culture system by co-culturing reticulocytes with endothelial cells and macrophages in a perfused 3D cell culture flow chamber, mimicking anatomy and physiology of a blood vessel in human microcirculation. In parallel, in vivo terminal maturation in an NSG mouse will be used to benchmark the key mechanistic steps and translate these to improve the 3D culture system. Directly comparing reticulocyte to erythrocyte transition of cultured red blood cells ex vivo to their physiologic maturation in vivo will for the first time allow us to monitor maturation over time and to characterize morphologic and functional changes in real time. Findings from our proposal will aid overcoming the hurdle towards clinical application of ex vivo expanded RBCs.