title: A hyperactive Mpl-based cell growth switch drives macrophage-associated erythropoiesis through an erythroid-megakaryocytic precursor.
abstract: Several approaches for controlling hematopoietic stem and progenitor cell expansion, lineage commitment, and maturation have been investigated for improving clinical interventions. We report here that amino acid substitutions in a thrombopoietin receptor (Mpl)--containing cell growth switch (CGS) extending receptor stability improve the expansion capacity of human cord blood CD34(+) cells in the absence of exogenous cytokines. Activation of this CGS with a chemical inducer of dimerization (CID) expands total cells 99-fold, erythrocytes 70-fold, megakaryocytes 0.5-fold, and CD34(+) stem/progenitor cells 4.4-fold by 21 days of culture. Analysis of cells in these expanded populations identified a CID-dependent bipotent erythrocyte-megakaryocyte precursor (PEM) population, and a CID-independent macrophage population. The CD235a(+)/CD41a(+) PEM population constitutes up to 13% of the expansion cultures, can differentiate into erythrocytes or megakaryocytes, exhibits very little expansion capacity, and exists at very low levels in unexpanded cord blood. The CD206(+) macrophage population constitutes up to 15% of the expansion cultures, exhibits high-expansion capacity, and is physically associated with differentiating erythroblasts. Taken together, these studies describe a fundamental enhancement of the CGS expansion platform, identify a novel precursor population in the erythroid/megakaryocytic differentiation pathway of humans, and implicate an erythropoietin-independent, macrophage-associated pathway supporting terminal erythropoiesis in this expansion system.
authors.affiliation: Department of Medicine, Division of Medical Genetics and Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA; Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, WA; and.
authors.forename: Eyayu
authors.initials: E
authors.lastname: Belay
authors.affiliation: Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA; Department of Medicine, Division of Hematology, University of Washington, Seattle, WA.
authors.forename: Chris P
authors.initials: CP
authors.lastname: Miller
authors.affiliation: Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA; Department of Medicine, Division of Hematology, University of Washington, Seattle, WA.
authors.forename: Amanda N
authors.initials: AN
authors.lastname: Kortum
authors.affiliation: Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, WA; and.
authors.forename: Beverly
authors.initials: B
authors.lastname: Torok-Storb
authors.affiliation: Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA; Department of Medicine, Division of Hematology, University of Washington, Seattle, WA.
authors.forename: C Anthony
authors.initials: CA
authors.lastname: Blau
authors.affiliation: Department of Medicine, Division of Medical Genetics and Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA;
authors.forename: David W
authors.initials: DW
authors.lastname: Emery
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D018952:Antigens, CD34 / Q000032:analysis; D002460:Cell Line; D049109:Cell Proliferation; D002478:Cells, Cultured; D041905:Erythroid Cells / Q000166:cytology* / Q000378:metabolism; D004920:Erythropoiesis; D005312:Fetal Blood / Q000166:cytology; D006801:Humans; D008533:Megakaryocytes / Q000166:cytology* / Q000378:metabolism; D051379:Mice; D040201:Platelet Membrane Glycoprotein IIb / Q000032:analysis; D053628:Receptors, Thrombopoietin / Q000235:genetics* / Q000378:metabolism
chemical_list: D018952:Antigens, CD34; D040201:Platelet Membrane Glycoprotein IIb; D053628:Receptors, Thrombopoietin; C082218:MPL protein, human