The Role of Hepatic Leukemia Factor in Hematopoiesis

Abstract: The development of therapeutic methods based on stem cells (e.g. cell therapy, activating regeneration and organ transplantation), known as regenerative medicine, is still a novel area in the field of life sciences. If successful, such therapies could be applicable to many diseases currently lacking effective treatments, such as Parkinson’s disease, diabetes, heart disease or cancer.Stem cells are cells with a unique potential to self-renew, that is divide into two identical daughter cells having the same capacity as the mother cell, and the potential to differentiate into mature cells. Adult (somatic) stem cells have been identified in many organs such as blood, skin and intestines in adult humans. These stem cells are important for the internal repair system due to their ability to divide essentially without limits to replace the short-lived mature cells. In this way, blood cell production is sustained throughout an individual’s lifetime by a few rare hematopoietic stem cells (HSC) residing in the bone marrow. Blood is a crucial part of the human body since blood cells provide oxygen and nutrients and protect the body against infections. Most of these cells are short-lived and are continuously recreated from HSCs through a gradual process of maturation. Furthermore, HSCs have the capacity to migrate from the bone marrow out to peripheral blood, a process termed mobilization, and to home back to their bone marrow microenvironment called niche. Due to these abilities, transplantations based on stem cell therapy are today routine clinical procedures as a life-saving treatment in many serious hematopoietic diseases such as leukemia and genetic blood disorders. However, not all patients can be treated in this way due to low HSC yields and the lack of matching donors. Therefore, a long-standing goal for the field has been to establish protocols that would allow for the expansion of HSCs in culture. However, efforts to expand self-renewing HSCs that can engraft efficiently have not been successful as yet due to our limited understanding of their self-renewal mechanisms.In steady state hematopoiesis, the majority of HSCs are in a dormant, non-dividing, quiescent state residing in the bone marrow niche. However, at each round of cell division, only a subset of stem cells becomes active, suggesting that different states of stem cells coexist within the same niche. We hypothesize that genetic modification might influence the hematopoietic stem cell function during homeostasis, regeneration, aging and leukemia. As we and others have previously demonstrated, the transcription factor, hepatic leukemia factor (HLF), is highly expressed in HSCs, the aim of this thesis is to define the role of HLF in HSC regulation using genetically modified mice lacking the HLF gene. In Article I, we confirmed that HLF is specifically expressed in the hematopoietic stem and progenitor cells and subsequently downregulated during differentiation. We show that HSCs devoid of Hlf display increased cell cycle and decreased HSC quiescence. In line, Hlf deficient HSCs demonstrated a poor ability to reconstitute hematopoiesis in serial transplantation assay, indicating a reduced self-renewal capacity. Furthermore, Hlf deficient mice were hypersensitive to chemotherapeutic agents, such as 5-FU, that completely eradicated the HSC pool, and Hlf deficient mice failed to recover following treatment. Interestingly, both by immunophenotype and on a transcriptional level, the Hlf deficient HSCs resembled more active short-term HSCs than long-term HSCs. Our findings unravel a novel role for HLF as a master regulator of HSC activity, highlighting the important role for HLF in hematopoiesis.Aging is a general important subject for all of us, and even more evident for people who are exposed to extremely high radiation levels, e.g. cancer patients undergoing chemotherapy or astronauts who work in space. Aging is correlated with reduced cellular fitness, which is seen in a decline in somatic stem cells functions. As such, aged HSCs are associated with altered lineage commitment and a reduced capacity to recreate the hematopoietic system upon transplantation. Still, HSCs are protected by intrinsic mechanisms to ensure a functional stem cell pool over time. Interestingly, our data show that HLF is expressed in the HSC compartment throughout ontogeny, and since we demonstrated in Article I that HLF is crucial to maintain the HSC pool during regenerations, subsequently we studied the role of HLF in aged HSCs and development. In Article II, we showed that, although important for maintaining the HSC pool during regeneration, HLF is dispensable for normal aging, indicating that the mechanism controlling aging differs from the mechanism regulating reconstitution after transplantation.Another key component of HSC regulation is their ability to home to the niche. In Article III, we developed a novel, standardized adhesion assay to investigate the interaction between HSCs and the niche represented by stroma cells. By using an RNA interference screen, we identified that Cytohesin 1 is a critical mediator of the attraction between HSCs and the stroma.