Generation Of Functional Erythrocytes From Human Embryonic Stem Cell-Derived Definitive Hematopoiesis

Division of Cellular Therapy, Advanced Clinical Research Center, Institute of Medical Science, University of Tokyo, Tokyo 108-8639, Japan.
Proceedings of the National Academy of Sciences (Impact Factor: 9.67). 10/2008; 105(35):13087-92. DOI: 10.1073/pnas.0802220105
Source: PubMed


A critical issue for clinical utilization of human ES cells (hESCs) is whether they can generate terminally mature progenies with normal function. We recently developed a method for efficient production of hematopoietic progenitors from hESCs by coculture with murine fetal liver-derived stromal cells. Large numbers of hESCs-derived erythroid progenitors generated by the coculture enabled us to analyze the development of erythropoiesis at a clone level and investigate their function. The results showed that the globin expression in the erythroid cells in individual clones changed in a time-dependent manner. In particular, embryonic epsilon-globin-expressing erythroid cells from individual clones decreased, whereas adult-type beta-globin-expressing cells increased to approximately 100% in all clones we examined, indicating that the cells undergo definitive hematopoiesis. Enucleated erythrocytes also appeared among the clonal progeny. A comparison analysis showed that hESC-derived erythroid cells took a similar differentiation pathway to human cord blood CD34(+) progenitor-derived cells when examined for the expression of glycophorin A, CD71 and CD81. Furthermore, these hESC-derived erythroid cells could function as oxygen carriers and had a sufficient glucose-6-phosphate dehydrogenase activity. The present study should provide an experimental model for exploring early development of human erythropoiesis and hemoglobin switching and may help in the discovery of drugs for hereditary diseases in erythrocyte development.

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Available from: Hiromi Sakai, Jun 11, 2014
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    • "Several groups showed that erythroblasts could be generated from ESCs, but the RBC yield was very low.28,29 In an industrial trial, Advanced Cell Technology (Worcester, MA, USA) first reported that ESCs differentiated into functional oxygen-carrying erythrocytes on a large scale (1010~1011 cells/6-well plate hESCs) as a source for clinical grade mass production of RBCs from stem cells.30 "
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    ABSTRACT: To date, the use of red blood cells (RBCs) produced from stem cells in vitro has not proved practical for routine transfusion. However, the perpetual and widespread shortage of blood products, problems related to transfusion-transmitted infections, and new emerging pathogens elicit an increasing demand for artificial blood. Worldwide efforts to achieve the goal of RBC production through stem cell research have received vast attention; however, problems with large-scale production and cost effectiveness have yet to prove practical usefulness. Some progress has been made, though, as cord blood stem cells and embryonic stem cells have shown an ability to differentiate and proliferate, and induced pluripotent stem cells have been shown to be an unlimited source for RBC production. However, transfusion of stem cell-derived RBCs still presents a number of challenges to overcome. This paper will summarize an up to date account of research and advances in stem cell-derived RBCs, delineate our laboratory protocol in producing RBCs from cord blood, and introduce the technological developments and limitations to current RBC production practices.
    Yonsei medical journal 03/2014; 55(2):304-9. DOI:10.3349/ymj.2014.55.2.304 · 1.29 Impact Factor
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    • "It was once expected that CD34 + cells from cord blood (CB) or bone marrow would someday provide a means for ex vivo expansion or in vitro generation of erythrocytes (red blood cells; RBCs) for transfusion (Giarratana et al., 2011), but the inability to produce sufficient numbers of CD34 + cells remains a bottleneck in this process . It was also thought that the advent of pluripotent stem cells (PSCs), including embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), would eliminate the need for blood donation, but a series of differentiation trials to create various blood cells from human ESCs or iPSCs has highlighted the difficulty of obtaining blood cells in quantities sufficient for use in transfusion using this method (Lu et al., 2007, 2011; Takayama et al., 2008, 2010). "
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    ABSTRACT: The lack of knowledge about the mechanism of erythrocyte biogenesis through self-replication makes the in vitro generation of large quantities of cells difficult. We show that transduction of c-MYC and BCL-XL into multipotent hematopoietic progenitor cells derived from pluripotent stem cells and gene overexpression enable sustained exponential self-replication of glycophorin A(+) erythroblasts, which we term immortalized erythrocyte progenitor cells (imERYPCs). In an inducible expression system, turning off the overexpression of c-MYC and BCL-XL enabled imERYPCs to mature with chromatin condensation and reduced cell size, hemoglobin synthesis, downregulation of GCN5, upregulation of GATA1, and endogenous BCL-XL and RAF1, all of which appeared to recapitulate normal erythropoiesis. imERYPCs mostly displayed fetal-type hemoglobin and normal oxygen dissociation in vitro and circulation in immunodeficient mice following transfusion. Using critical factors to induce imERYPCs provides a model of erythrocyte biogenesis that could potentially contribute to a stable supply of erythrocytes for donor-independent transfusion.
    Stem Cell Reports 12/2013; 1(6):499-508. DOI:10.1016/j.stemcr.2013.10.010 · 5.37 Impact Factor
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    • "In addition to shedding light on the developmental biology of erythroid cells, the investigation of HESC-derived erythroblasts (ESERs) and/or HiPSC-derived erythroid cells may be clinically relevant. ESERs and HiPSC-derived erythroblasts reportedly exhibit characteristics of cord blood (CB) in terms of their oxygen dissociation curve, G6PD activities , CO rebinding kinetics, and response to 2,3-DPG depletion [7] [8] [9]. Cell enucleation can also occur under appropriate culture conditions [5]. "
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    ABSTRACT: To explore the mechanisms controlling erythroid differentiation and development, we analyzed the genome-wide transcription dynamics occurring during the differentiation of human embryonic stem cells (HESCs) into the erythroid lineage and development of embryonic to adult erythropoiesis using high throughput sequencing technology. HESCs and erythroid cells at three developmental stages: ESER (embryonic), FLER (fetal), and PBER (adult) were analyzed. Our findings revealed that the number of expressed genes decreased during differentiation, whereas the total expression intensity increased. At each of the three transitions (HESCs-ESERs, ESERs-FLERs, and FLERs-PBERs), many differentially expressed genes were observed, which were involved in maintaining pluripotency, early erythroid specification, rapid cell growth, and cell-cell adhesion and interaction. We also discovered dynamic networks and their central nodes in each transition. Our study provides a fundamental basis for further investigation of erythroid differentiation and development, and has implications in using ESERs for transfusion product in clinical settings.
    Genomics 10/2013; 102(5-6). DOI:10.1016/j.ygeno.2013.09.005 · 2.28 Impact Factor
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