Expression and role of the cytokine tyrosine kinase receptor flt3 in early hematopoiesis

Sammanfattning: Mature blood cells are crucial for life, but they have a short lifetime and thus have to be replaced. These mature blood cells are produced by lineage restricted progenitors, which themselves are generated by rare multipotent hematopoietic stem cells (HSCs) in a highly dynamic process called hematopoiesis. In addition to the ability of HSCs to generate all blood cell lineages, they possess the unique property of self-renewal, a process in which a HSC, during a cell division generate at least one daughter cell identical to the parental cell. The maintenance and regulation of HSCs and earliest stages of lineage development are partly controlled by soluble and membrane bound regulators called cytokines. The focus in this thesis has been on the cytokine tyrosine kinase receptor flt3 and its ligand and their role in regulating HSCs and the earliest progenitors in adult murine bone marrow (BM). Earlier studies had implicated a role for flt3 and its ligand in the maintenance of HSCs. When studying mice with targeted deletion of the flt3 ligand (FL), we failed to find any role of flt3 and its ligand in HSC regulation. However, the generation of the common lymphoid progenitor (CLP) and earliest stages of B- and T cell development were severely affected, but not myeloid progenitors or later stages of lymphoid development. Based on flt3 expression, we subfractionated the Lin(-)Sca-1(+)c-kit(+) (LSK) stem cell pool in mouse bone marrow. In contrast to LSKflt3- cells, which sustained multilineage long-term reconstitution, LSKflt3+ cells generated only short-term lymphoid dominated reconstitution when injected into lethally ablated recipients. We also demonstrated the hierarchical relationship at the earliest stages of hematopoiesis, in that LSKflt3- HSCs generate LSKflt3+ cells, which generate the CLP but not the LSKflt3- cells. In the classical model of the hematopoietic hierarchy, the first lineage commitment step of HSCs results in a strict separation into distinct lymphoid and myeloid pathways, generating the recently identified CLP and the common myeloid progenitor (CMP). However, in sharp contrast to LSKflt3-, cells which could generate all blood cell lineages, LSK cells expressing flt3 could not generate megakaryocytes or erythroid cells. Thus, these finding together with the above mentioned relationship between the LSKflt3-, LSKflt3+ and CLP do not support the classical hematopoietic hierarchy model depicting the first lineage commitment generating a strict separation of lymphoid and myeloid pathways. The LSK population contains all LT-HSC activity. However, this population is not homogenous neither in phenotype or function and it has been proposed to contain at least two populations, one with long-term reconstitution (LTR) potential and one with short-term reconstitution potential. Based on the expression of CD34 and flt3 within the adult LSK stem cell pool we were able to subfractionate short-term hematopoietic stem cells (ST-HSC) from the LT-HSCs. In contrast to the LSKCD34-flt3- and LSKCD34+flt3+ cells, the LSKCD34+flt3- is highly enriched for CFU-S activity and capable of rescuing lethally irradiated recipients, fulfilling the criteria of ST-HSCs.