title: Clonal analysis of hematopoietic progenitor cells in the zebrafish.
abstract: Identification of hematopoietic progenitor cells in the zebrafish (Danio rerio) has been hindered by a lack of functional assays to gauge proliferative potential and differentiation capacity. To investigate the nature of myeloerythroid progenitor cells, we developed clonal methylcellulose assays by using recombinant zebrafish erythropoietin and granulocyte colony-stimulating factor. From adult whole kidney marrow, erythropoietin was required to support erythroid colony formation, and granulocyte colony-stimulating factor was required to support the formation of colonies containing neutrophils, monocytes, and macrophages. Myeloid and erythroid colonies showed distinct morphologies and were easily visualized and scored by their expression of lineage-specific fluorescent transgenes. Analysis of the gene-expression profiles after isolation of colonies marked by gata1:DsRed or mpx:eGFP transgenes confirmed our morphological erythroid and myeloid lineage designations, respectively. The majority of progenitor activity was contained within the precursor light scatter fraction, and more immature precursors were present within the lymphoid fraction. Finally, we performed kinetic analyses of progenitor activity after sublethal irradiation and demonstrated that recovery to preirradiation levels occurred by 14 days after irradiation. Together, these experiments provide the first report of clonal hematopoietic progenitor assays in the zebrafish and establish the number, characteristics, and kinetics of myeloerythroid progenitors during both steady-state and stress hematopoiesis.
authors.affiliation: Department of Cellular and Molecular Medicine, University of California at San Diego, La Jolla, CA 92093-0380, USA.
authors.forename: David L
authors.initials: DL
authors.lastname: Stachura
authors.forename: Ondrej
authors.initials: O
authors.lastname: Svoboda
authors.forename: Ryan P
authors.initials: RP
authors.lastname: Lau
authors.forename: Keir M
authors.initials: KM
authors.lastname: Balla
authors.forename: Leonard I
authors.initials: LI
authors.lastname: Zon
authors.forename: Petr
authors.initials: P
authors.lastname: Bartunek
authors.forename: David
authors.initials: D
authors.lastname: Traver
mesh_terms: D000818:Animals; D030801:Animals, Genetically Modified; D002454:Cell Differentiation / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D002999:Clone Cells; D004625:Embryo, Nonmammalian; D041905:Erythroid Cells / Q000187:drug effects; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006410:Hematopoiesis / Q000187:drug effects / Q000235:genetics / Q000502:physiology; D006412:Hematopoietic Stem Cells / Q000166:cytology* / Q000187:drug effects / Q000502:physiology*; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology; D022423:Myeloid Cells / Q000187:drug effects / Q000502:physiology; D011994:Recombinant Proteins / Q000494:pharmacology; D015027:Zebrafish / Q000196:embryology* / Q000235:genetics / Q000502:physiology
chemical_list: D036341:Intercellular Signaling Peptides and Proteins; D011994:Recombinant Proteins; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins