Towards Generation of Human Hematopoietic Stem Cells from Pluripotent Stem Cells

Abstract: There is a pressing need for compatible Hematopoietic Stem Cells (HSCs), in high quantities and of good clinical quality, to ensure optimal treatment for patients with various blood malignancies. Pluripotent Stem Cells (PSCs) have the potential to be a source for generating HSCs. The indefinite expansion capacity of PSCs, combined with the possibility of creating these cells from patient-specific material could, in principle, allow for the generation of patient-derived HSCs. Generation of HSCs from PSCs could dramatically improve the treatment of patients in the clinic, including the possible correction of genetically carried hematological diseases. Furthermore, successful generation of HSCs would provide a tool to better study the nature of hematological malignancies, and could be employed for the purpose of drug screening With the ultimate goal of facilitating in vitro generation of functional HSCs from human PSCs, the studies included in this thesis explore potential key factors and mechanisms involved in the process of hematopoietic development. In paper I we describe that control over the amount of retinoic acid during in vitro differentiation of human PSCs increase blood generation, allowing for more defined and efficient production of relevant cells. In paper II we provide a molecular identification of Endothelial to Hematopoietic Transition (EHT), the conversion through which HSCs emerge from endothelial cells. This provides a foundation for further characterization of the cells undergoing this transition, and could potentially enable control over this process for the purpose of more efficient generation of hematopoietic cells from hemogenic endothelium. In paper III we explore how cAMP intrinsically regulates the generation of HSC-like cells, and we demonstrate the importance of cAMP signaling in the emergence of HSCs. Modulating cAMP provides an additional approach to increase the generation, and potentially the function, of HSC-like cells from human PSCs. In paper IV, we demonstrate that reactive oxygen species is increased due to in vitro culture, causing functional impairment of hematopoietic progenitors and HSC-like cells generated from human PSCs. This argues that we must revise our standardized culture methods, and that a more physiological oxygen concentration is likely to be a critical component for handling and generating HSCs outside of the body. In summary, the papers included in this thesis focuses on understanding the development of human HSCs, with the ultimate goal of developing methods that would enable the generation of functional HSCs in the laboratory, potentially translating into improved treatments against a multitude of hematological malignancies.