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Abstract

Hematopoietic stem cells (HSCs) that reside in the bone marrow are known to be of vital importance in generating diverse peripheral blood cell types, both under homeostasis and stress-hematopoiesis. Stressors such as infections and inflammatory cytokines such as IFNα can have direct effects on HSC biology. This exemplifies the importance of these factors in stress-hematopoiesis, and hematological neoplasms driven by mutant HSCs. To this end, two different aspects of these factors were studied in this dissertation – The mechanism of action of IFNα as a therapeutic agent in the treatment of myeloproliferative neoplasm (MPN), and co-housing as a murine model system to study the effect of infections on HSCs. The mechanism of action of pegylated-IFNα (peg-IFNα) in reducing mutant allelic burden in JAK2-mutated MPNs was studied, in comparison to JAK inhibitor therapy. Using a BrdU pulse-chase assay in a mouse model of MPN, it was observed that JAK2-mutant HSCs have an inherently higher proliferation compared to wildtype-HSCs. However, a sub-fraction of mutant HSCs was found to persist in a dormant state in the bone marrow. These dormant mutant HSCs may be responsible for disease propagation and offer resistance to therapy with JAK inhibitors such as Fedratinib. In support of this, dormant JAK2-mutant HSCs were unaffected by Fedratinib treatment in mice, where hematologic parameters had been normalized. On the other hand, peg-IFNα was found to eliminate the dormant mutant HSCs in concomitance with reduced mutant-allele burden. These data hence provide a potential mechanism of action of peg-IFNα in treating MPNs. Chronic infections in mice have been shown to lead to an increased proliferation of the hematopoietic stem and progenitor cell (HSPC) compartment and a subsequent loss of stem cell function. However, many of these infection-models involve challenging mice with a very high dose of purified pathogen administered via a non-physiologic route, thereby not recapitulating a natural infection process. To this end, a co-housing model was assessed, which allows natural transmission of infections from so-called “wildling” mice, to specific-pathogen-free laboratory mice, termed “co-housed” mice. The wildling mice have a microbiota and pathogen spectrum, similar to that of a wild mouse, which they aquire by neonatal vertical transmission. The co-housing model was observed to have an efficient horizontal transmission of multiple infections from wildling to co-housed mice. Both acute and chronic phases of co-housing mediated infections were observed to elevate the peripheral blood cell count in the co-housed mice, but did not lead to expansion of HSPCs in the bone marrow. The chronic infections acquired through co-housing were found to reduce the functional potency of HSCs in the co-housed mice. This was in line with observations in previous infection models studied by other groups, albeit to a milder degree. Furthermore, the effect of vertical transmission was studied in the wildling mice. It was found that the HSCs from these mice did not have any reduction in function, contrary to what was observed in the co-housed mice. This indicates that neonatal exposure to pathogens preserves HSC function, suggesting a context-specific window of opportunity for infection-mediated functional attrition of HSCs. Overall, the findings from this doctoral dissertation have contributed to the field of hematology from two unique perspectives. This work has provided a new paradigm to further explore novel strategies to eliminate therapy resistant HSCs in the setting of JAK2 mutated MPNs. It has also led to the development of a model to study differential response of HSCs to infections depending on the timing of exposure to pathogens. Together, these contributions can drive development of new therapeutic options for MPN patients, and also further our understanding of the roles of infections in hematopoiesis and hematologic malignancies.