Elevated Functional Hematopoietic Stem and Progenitor Cells in the Peripheral Blood of G-CSF-Mobilized Dnmt3a-Mutant Mice

Abstract

Clonal hematopoiesis (CH) arising from somatic mutations in hematopoietic stem cells (HSCs) is associated with increased risk of developing several non-hematologic and hematologic diseases. In the context of stem cell transplantation, donor HSCs carrying CH-associated somatic mutations can be passed from donor to recipient during stem cell transplantation. Most literature supports that there is no negative impact on recipient survival or outcomes after allogeneic stem cell transplantation (allo-HSCT) from donors with CH. However, it is less clear how allo-HSCT donors with CH respond to common mobilization methods such as Neupogen (G-CSF). As hematopoietic stem and progenitor cells (HSPCs) with CH mutations have been found to respond to external stimuli such as inflammation in a manner distinct from wild-type HSPCs, it is plausible that CH-mutant HSPCs also distinctly respond to G-CSF mobilization. It is important to understand whether the process of mobilization differentially enriches for CH-mutant HSPCs in the mobilized peripheral blood product. To address this gap in knowledge, we used a murine model of Dnmt3a-mutant CH. After five consecutive days of Neupogen (G-CSF) administration, modeling the procedure for human HSC mobilization, we observed that Dnmt3a-mutant mice had significantly increased white blood cell, neutrophil and monocyte counts compared to control mice. We observed a significant increase in cell surface marker-defined HSPCs in the peripheral blood and spleen of G-CSF-treated Dnmt3a-mutant mice compared to G-CSF-treated control mice. Using in vitro colony-forming unit assays, the peripheral blood of G-CSF-treated Dnmt3a-mutant mice was found to contain a greater number of functional hematopoietic progenitors compared to G-CSF-treated control mice. To determine if the peripheral blood of G-CSF-treated Dnmt3a-mutant mice had a greater abundance of functional HSCs, we transplanted equal volumes of mobilized blood from Dnmt3a-mutant and control mice into primary and secondary lethally irradiated wild-type recipient mice. We observed increased long-term engraftment of mobilized blood from Dnmt3a-mutant mice compared to control mice in both primary and secondary recipient animals. At a mechanistic level, HSC mobilization is facilitated by myeloid cells that respond directly to G-CSF via the G-CSF receptor ( Csf1r). In Dnmt3a-mutant mice, we observed increased Csf1r expression in myeloid progenitor and mature myeloid cells compared to control mice, providing a putative mechanism by which increased mobilization is enabled. Together, our data support differential mobilization potential of Dnmt3a-mutant compared to control HSCs in response to G-CSF, which may have implications in the context of allo-HSCT.