scholarly journals Globin switches in yolk sac–like primitive and fetal-like definitive red blood cells produced from human embryonic stem cells

Blood ◽  
2008 ◽  
Vol 111 (4) ◽  
pp. 2400-2408 ◽  
Author(s):  
Caihong Qiu ◽  
Emmanuel N. Olivier ◽  
Michelle Velho ◽  
Eric E. Bouhassira
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Fetal Liver ◽  
Embryonic Stem ◽  
Fetal Hemoglobin ◽  
Yolk Sac ◽  
Globin Mrna ◽  
Fetal Hepatocytes ◽  
Cd34 Cells

We have previously shown that coculture of human embryonic stem cells (hESCs) for 14 days with immortalized fetal hepatocytes yields CD34+ cells that can be expanded in serum-free liquid culture into large numbers of megaloblastic nucleated erythroblasts resembling yolk sac–derived cells. We show here that these primitive erythroblasts undergo a switch in hemoglobin (Hb) composition during late terminal erythroid maturation with the basophilic erythroblasts expressing predominantly Hb Gower I (ζ2ϵ2) and the orthochromatic erythroblasts hemoglobin Gower II (α2ϵ2). This suggests that the switch from Hb Gower I to Hb Gower II, the first hemoglobin switch in humans is a maturation switch not a lineage switch. We also show that extending the coculture of the hESCs with immortalized fetal hepatocytes to 35 days yields CD34+ cells that differentiate into more developmentally mature, fetal liver–like erythroblasts, that are smaller, express mostly fetal hemoglobin, and can enucleate. We conclude that hESC-derived erythropoiesis closely mimics early human development because the first 2 human hemoglobin switches are recapitulated, and because yolk sac–like and fetal liver–like cells are sequentially produced. Development of a method that yields erythroid cells with an adult phenotype remains necessary, because the most mature cells that can be produced with current systems express less than 2% adult β-globin mRNA.

scholarly journals Hematopoietic differentiation of human embryonic stem cells progresses through sequential hematoendothelial, primitive, and definitive stages resembling human yolk sac development

Blood ◽  
2005 ◽  
Vol 106 (3) ◽  
pp. 860-870 ◽  
Author(s):  
Elias T. Zambidis ◽  
Bruno Peault ◽  
Tea Soon Park ◽  
Fred Bunz ◽  
Curt I. Civin
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Embryonic Stem ◽  
Homeobox Gene ◽  
Fetal Hemoglobin ◽  
Yolk Sac ◽  
Embryoid Bodies ◽  
Hematopoietic Differentiation ◽  
Hematopoietic Stem

AbstractWe elucidate the cellular and molecular kinetics of the stepwise differentiation of human embryonic stem cells (hESCs) to primitive and definitive erythromyelopoiesis from human embryoid bodies (hEBs) in serum-free clonogenic assays. Hematopoiesis initiates from CD45 hEB cells with emergence of semiadherent mesodermal-hematoendothelial (MHE) colonies that can generate endothelium and form organized, yolk sac–like structures that secondarily generate multipotent primitive hematopoietic stem progenitor cells (HSPCs), erythroblasts, and CD13+CD45+ macrophages. A first wave of hematopoiesis follows MHE colony emergence and is predominated by primitive erythropoiesis characterized by a brilliant red hemoglobinization, CD71/CD325a (glycophorin A) expression, and exclusively embryonic/fetal hemoglobin expression. A second wave of definitive-type erythroid burst-forming units (BFU-e's), erythroid colony-forming units (CFU-e's), granulocyte-macrophage colony-forming cells (GM-CFCs), and multilineage CFCs follows next from hEB progenitors. These stages of hematopoiesis proceed spontaneously from hEB-derived cells without requirement for supplemental growth factors during hEB differentiation. Gene expression analysis of differentiating hEBs revealed that initiation of hematopoiesis correlated with increased levels of SCL/TAL1, GATA1, GATA2, CD34, CD31, and the homeobox gene-regulating factor CDX4 These data indicate that hematopoietic differentiation of hESCs models the earliest events of embryonic and definitive hematopoiesis in a manner resembling human yolk sac development, thus providing a valuable tool for dissecting the earliest events in human HSPC genesis.

Monocytic cells derived from human embryonic stem cells and fetal liver share common differentiation pathways and homeostatic functions

Blood ◽  
2011 ◽  
Vol 117 (11) ◽  
pp. 3065-3075 ◽  
Author(s):  
Olena Klimchenko ◽  
Antonio Di Stefano ◽  
Birgit Geoerger ◽  
Sofiane Hamidi ◽  
Paule Opolon ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Fetal Liver ◽  
Critical Role ◽  
Embryonic Stem ◽  
Blood Monocytes ◽  
Monocytic Cell ◽  
Monocytic Cells

Abstract The early emergence of macrophages and their large pattern of tissue distribution during development suggest that they may play a critical role in the initial steps of embryogenesis. In the present study, we show that monocytic cells derived from human embryonic stem cells (hESCs) and from fetal liver follow a differentiation pathway different to that of adult cells, leading to specific functions. Embryonic and fetal monocytic cells differentiated from a CD14lowCD16− precursor to form CD14highCD16+ cells without producing the CD14highCD16− cell population that predominates in adult peripheral blood. Both demonstrated an enhanced expression of genes encoding tissue-degrading enzymes, chemokines, and scavenger receptors, as was previously reported for M2 macrophages. Compared with adult blood monocytes, embryonic and fetal monocytic cells secreted high amounts of proteins acting on tissue remodeling and angiogenesis, and most of them expressed the Tie2 receptor. Furthermore, they promoted vascular remodeling in xenotransplanted human tumors. These findings suggest that the regulation of human fetal and embryonic monocytic cell differentiation leads to the generation of cells endowed mainly with anti-inflammatory and remodeling functions. Trophic and immunosuppressive functions of M2-polarized macrophages link fetus and tumor development, and hESCs offer a valuable experimental model for in vitro studies of mechanisms sustaining these processes.

scholarly journals Definitive-like erythroid cells derived from human embryonic stem cells coexpress high levels of embryonic and fetal globins with little or no adult globin

Blood ◽  
2006 ◽  
Vol 108 (5) ◽  
pp. 1515-1523 ◽  
Author(s):  
Kai-Hsin Chang ◽  
Angelique M. Nelson ◽  
Hua Cao ◽  
Linlin Wang ◽  
Betty Nakamoto ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Molecular Mechanisms ◽  
Embryoid Body ◽  
Fetal Liver ◽  
Embryonic Stem ◽  
Erythroid Differentiation ◽  
Erythroid Cells ◽  
Promising Tool

Human embryonic stem cells are a promising tool to study events associated with the earliest ontogenetic stages of hematopoiesis. We describe the generation of erythroid cells from hES (H1) by subsequent processing of cells present at early and late stages of embryoid body (EB) differentiation. Kinetics of hematopoietic marker emergence suggest that CD45+ hematopoiesis peaks at late D14EB differentiation stages, although low-level CD45- erythroid differentiation can be seen before that stage. By morphologic criteria, hES-derived erythroid cells were of definitive type, but these cells both at mRNA and protein levels coexpressed high levels of embryonic (ϵ) and fetal (γ) globins, with little or no adult globin (β). This globin expression pattern was not altered by the presence or absence of fetal bovine serum, vascular endothelial growth factor, Flt3-L, or coculture with OP-9 during erythroid differentiation and was not culture time dependent. The coexpression of both embryonic and fetal globins by definitive-type erythroid cells does not faithfully mimic either yolk sac embryonic or their fetal liver counterparts. Nevertheless, the high frequency of erythroid cells coexpressing embryonic and fetal globin generated from embryonic stem cells can serve as an invaluable tool to further explore molecular mechanisms.

Cells Extract from Fetal Liver Promotes the Hematopoietic Differentiation of Human Embryonic Stem Cells

2009 ◽  
Vol 11 (1) ◽  
pp. 51-60 ◽  
Author(s):  
Yu-Xiao Liu ◽  
Lei Ji ◽  
Wen Yue ◽  
Zhi-Feng Yan ◽  
Jing Wang ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Fetal Liver ◽  
Embryonic Stem ◽  
Hematopoietic Differentiation

A Novel Method for Efficient Production of Multipotential Hematopoietic Progenitors from Human Embryonic Stem Cells by Co-Culture with Murine Fetal Liver-Derived Stromal Cells.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 4214-4214
Author(s):  
Feng Ma ◽  
Dan Wang ◽  
Sachiyo Hanada ◽  
Hirohide Kawasaki ◽  
Yuji Zaike ◽  
...  
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Stromal Cells ◽  
Blood Cells ◽  
Fetal Liver ◽  
Hematopoietic Cells ◽  
Embryonic Stem ◽  
Efficient Production ◽  
Hematopoietic Progenitors

Abstract Human embryonic stem cells provide a unique tool to study early events occurring in the development of human embryonic hematopoiesis, and their totipotent capability indicates a potent clinical application based on the cellular therapy and the evaluation of drug effects on hematopoietic and blood cells. To achieve efficient production of hematopoietic cells from human embryonic stem cells, we attempted to reproduce the circumstance surrounding embryonic hematopoietic cells in vitro. Since fetal liver is the predominant source of hematopoietic and blood cells in mammalian embryogenesis, we established stromal cells from mouse fetal liver at days 14 to 15 of gestation. In the co-culture of human embryonic stem cells with the established stromal cells, a number of hematopoietic progenitors were generated at around day 14 of co-culture, and this hematopoietic activity was highly enriched in the cobble stone-like cells under the stromal layer. Most of the cobble stone-like cells collected expressed CD34 and contained a variety of hematopoietic colony-forming cells, especially multilineage colony-forming cells, at a high frequency. The multipotential hematopoietic progenitors in the cobble stone-like cells produced all types of mature blood cells, including adult type hemoglobin-synthesizing erythrocytes and tryptase and chymase-bouble positive mast cells in the suspension cultiue with a cytokine cocktail. The developed co-culture system of human embryonic stem cells should offer a novel source for hematopoietic and blood cells applicable to cellular therapies and drug screening.

scholarly journals Distinct DNA methylation patterns characterize differentiated human embryonic stem cells and developing human fetal liver

2009 ◽  
Vol 19 (6) ◽  
pp. 1044-1056 ◽  
Author(s):  
A. L. Brunner ◽  
D. S. Johnson ◽  
S. W. Kim ◽  
A. Valouev ◽  
T. E. Reddy ◽  
...  
Keyword(s):  
Dna Methylation ◽  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Fetal Liver ◽  
Embryonic Stem ◽  
Human Fetal Liver ◽  
Methylation Patterns

Step-Wise Differentiation of CD34+ Cell Derived from Nuclear Transfer-Human Embryonic Stem Cells into Myeloid and Lymphoid Precursors.

Blood ◽  
2005 ◽  
Vol 106 (11) ◽  
pp. 3614-3614
Author(s):  
Gabsang Lee ◽  
Jae-hung Sheih ◽  
Woosuk Hwang ◽  
Malcolm A.S. Moore
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Human Embryonic Stem Cells ◽  
Nuclear Transfer ◽  
Embryonic Stem ◽  
Erythroid Cell ◽  
Cell Marker ◽  
Hematopoietic Stem ◽  
Cd34 Cells ◽  
Hesc Lines

Abstract Human embryonic stem cells derived after nuclear transfer (NT-hESCs) may be used to study the earliest events in human hematopoietic stem cell development. They may also be potential candidates for cell therapy in degenerative and genetic diseases because of immunological compatibility, important for eventual transplantation. We developed an efficient, step-wise system for differentiating hESC to CD34+ hematopoietic stem cells from embryoid bodies (EB) in liquid cultures and, subsequently, further differentiating these into myeloid and lymphoid precursors using different combinations of cytokines and stromal cells. The H1 and NT-hESC-1 lines were maintained on mouse embryonic fibroblasts and NT-hESC-2 was cultured on autologous human fibroblasts with basic fibroblast growth factor in serum-replacement containing medium. In all three lines initiation of hematopoiesis begins during the first week of EB differentiation in the presence of different combinations of hematopoietic growth factors and in the absence of stromal support. Gene expression analysis, using RT-PCR methods, revealed that the first wave of embryonic hematopoiesis at 4–8 days of EB development was associated with expression of SCL/TAL1, flt3, KDR and GATA2. In FACS analysis, CD34+, KDR+, CD31+ cells were firstly detected in 6–10 days and CD34+, KDR+, CD45+, CD14+ (monocyte/macrophage marker), CD33+ (early myeloid cell marker), CD56+ (natural killer cell marker), Glycoporin A+ (erythroid cell marker), CD86 + (dendritic cell marker) cells were observed up to 15–20 days of EB development. CD34+ cell derived from 10–12 day EB’s from all three hESC lines, formed multiple lineage colonies in methylcellulose culture. These CD34+ cells can be maintained on OP9- stroma transduced with an adenovector expressing thrombopoietin (Tpo). We developed selective culture conditions for differentiation of CD34+ cells into different lineages. Isolated CD34+ cells from 10 to 12 Day EB’s differentiated in CD86+ dendritic cells in the presence of SCF, Tpo, Flt3 ligand, G-CSF, and TNF-alpha. Mature multinucleated CD41a+ megakaryocytes developed in cultures with SCF, Flt3L, and Tpo. Glycophorin A+ erythroid differentiation through to enucleated red cell stage was obtained with SCF, Epo, and IGF-1. B cell precursors (CD19+) developed in MS-5 stromal co-culture with SCF and G-SCF while CD44+CD25+ (early T progenitor marker) cells were detected in co-culture with OP9 transduced with the Notch ligand delta like-1 and IL-7. We have developed GFP and luciferase-expressing H1 and NT-hES-2 lines by lentiviral-vector-mediated gene transfer. These lines will allow us to monitor ES-derived cell localization and quantitation in immunodeficient mice using whole animal luciferase imaging systems. We are examining the potential for GFP-luciferase-transduced NT-hES-derived hematopoietic cells to engraft in NOD/SCID beta2−/ − mice. Our data demonstrate that NT-hESC lines as well as H1 lines are capable of differentiation to various lymphoid and hematopoietic lineages.

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