title: Extensive ex vivo expansion of functional human erythroid precursors established from umbilical cord blood cells by defined factors.
abstract: There is a constant shortage of red blood cells (RBCs) from sufficiently matched donors for patients who need chronic transfusion. Ex vivo expansion and maturation of human erythroid precursors (erythroblasts) from the patients or optimally matched donors could represent a potential solution. Proliferating erythroblasts can be expanded from umbilical cord blood mononuclear cells (CB MNCs) ex vivo for 10(6)-10(7)-fold (in ~50 days) before proliferation arrest and reaching sufficient number for broad application. Here, we report that ectopic expression of three genetic factors (Sox2, c-Myc, and an shRNA against TP53 gene) associated with iPSC derivation enables CB-derived erythroblasts to undergo extended expansion (~10(68)-fold in ~12 months) in a serum-free culture condition without change of cell identity or function. These expanding erythroblasts maintain immature erythroblast phenotypes and morphology, a normal diploid karyotype and dependence on a specific combination of growth factors for proliferation throughout expansion period. When being switched to a terminal differentiation condition, these immortalized erythroblasts gradually exit cell cycle, decrease cell size, accumulate hemoglobin, condense nuclei and eventually give rise to enucleated hemoglobin-containing erythrocytes that can bind and release oxygen. Our result may ultimately lead to an alternative approach to generate unlimited numbers of RBCs for personalized transfusion medicine.
authors.affiliation: Division of Hematology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; Stem Cell Program, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
authors.forename: Xiaosong
authors.initials: X
authors.lastname: Huang
authors.affiliation: Division of Hematology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; Stem Cell Program, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
authors.forename: Siddharth
authors.initials: S
authors.lastname: Shah
authors.affiliation: Division of Hematology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; Stem Cell Program, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
authors.forename: Jing
authors.initials: J
authors.lastname: Wang
authors.affiliation: Division of Hematology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; Stem Cell Program, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
authors.forename: Zhaohui
authors.initials: Z
authors.lastname: Ye
authors.affiliation: Division of Hematology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; Stem Cell Program, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
authors.forename: Sarah N
authors.initials: SN
authors.lastname: Dowey
authors.affiliation: Division of Hematology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; Stem Cell Program, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
authors.forename: Kit Man
authors.initials: KM
authors.lastname: Tsang
authors.affiliation: Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Laurel G
authors.initials: LG
authors.lastname: Mendelsohn
authors.affiliation: Hematology Branch, National Heart, Lung and Blood Institute, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Gregory J
authors.initials: GJ
authors.lastname: Kato
authors.affiliation: Division of Hematology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA.
authors.forename: Thomas S
authors.initials: TS
authors.lastname: Kickler
authors.affiliation: Division of Hematology, Department of Medicine, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA; Stem Cell Program, Institute for Cell Engineering, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA. Electronic address: lcheng2@jhmi.edu.
authors.forename: Linzhao
authors.initials: L
authors.lastname: Cheng
mesh_terms: D002454:Cell Differentiation / Q000235:genetics; D049109:Cell Proliferation; D016000:Cluster Analysis; D016207:Cytokines / Q000378:metabolism / Q000494:pharmacology; D004171:Diploidy; D004900:Erythroblasts / Q000166:cytology / Q000378:metabolism; D015672:Erythroid Precursor Cells / Q000166:cytology* / Q000378:metabolism*; D004920:Erythropoiesis / Q000502:physiology*; D005312:Fetal Blood / Q000166:cytology*; D020869:Gene Expression Profiling; D005786:Gene Expression Regulation; D020570:Genes, myb; D016259:Genes, myc; D006454:Hemoglobins / Q000235:genetics / Q000378:metabolism; D006728:Hormones / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D059785:Karyotype; D055748:SOXB1 Transcription Factors / Q000235:genetics
chemical_list: D016207:Cytokines; D006454:Hemoglobins; D006728:Hormones; C527782:SOX2 protein, human; D055748:SOXB1 Transcription Factors