Erythropoietin Signaling Inhibits Long Term Marrow Reconstitution

Murine hematopoietic stem cells (HSC) transfected with a gain-of-function human erythropoietic receptor (EPOR) transgene were reported to have a competitive advantage over wild type mouse hematopoietic stem cells in a bone marrow transplantation (BMT) model (Kirby, Blood 95(12): 3710, 2000). However, EPOR transgenes may not be normally expressed in early progenitor/stem cells. Moreover, whether Epo/EpoR signaling plays a role in hematopoietic stem cell engraftment is unknown. Our lab previously created mouse models harboring the wild type human EPOR (wthEPOR) or the mutant human gain-of-function EPOR (mthEPOR) gene knocked into the mouse EPOR locus (Divoky, PNAS 98(3): 986, 2001). This animal model has augmented Epo signaling in all tissues that express EpoR, thus the wthEPOR mice are anemic while the mthEPOR mice are polycythemic. We compared the relative engraftment efficiency of mthEPOR vs. wthEPOR HSCs in a competitive bone marrow transplantation (BMT) assay using C57/Bl6 congenic mice. Bone marrow from wthEPOR (CD45.1) and mthEPOR (CD45.2) mice were co-transplanted (1:1) into lethally irradiated (137Cs > 11Gy split) normal recipients (CD45.1/CD45.2). At 7 months after transplantation, peripheral blood chimerism demonstrated skewing towards wthEPOR rather than mthEPOR origin in the granulocyte, macrophage, T cell, and B cell compartments (Data Table). Bone marrow chimerism paralleled peripheral blood chimerism (not shown). Examination of the stem cell compartment by Hoechst 33342 staining demonstrated similar skewing towards wthEPOR origin (Data Table).
Because unequal numbers of HSC may result in skewed chimerism, we examined the relative proportions of HSC to total bone marrow cells. In wthEPOR mice, the Flt3- Rh123^low subset of cKit⁺Sca¹+ cells (KLS-FS) cells represented 0.011±0.003% of total bone marrow cells while in mthEPOR mice these cells represented 0.023±0.006% of total bone marrow cells (p=0.025). Since equal numbers of wthEPOR and mthEPOR total bone marrow cells were co-transplanted, relatively fewer wthEPOR HSC than mthEPOR HSC were transferred. Taken with the above chimerism data showing skewing towards wthEPOR, these results suggest that wthEPOR HSCs have a significant engraftment advantage over mthEPOR HSCs. Furthermore, enhanced Epo/EpoR signaling may interfere with the long term repopulation of hematopoietic progenitors. Hematopoietic stem cells undergo self renewal or differentiation/proliferation; in the presence of erythropoietin, a cytokine with proliferative and differentiating properties, it may be that self renewal is suppressed leading ultimately to the observed skewed chimerism. These data suggest that erythropoietin administration to patients during and immediately after marrow transplantation may be detrimental and should be used judiciously.