In vitro human embryonic stem cell hematopoiesis mimics MYB-independent yolk sac hematopoiesis
Stijn Vanhee,
Katrien De Mulder,
Yasmine Van Caeneghem,
Greet Verstichel,
Nadine Van Roy,
Björn Menten,
Imke Velghe,
Jan Philippé,
Dominique De Bleser,
Bart N. Lambrecht,
Tom Taghon,
Georges Leclercq,
Tessa Kerre,
Bart Vandekerckhove
Affiliations
Stijn Vanhee
Laboratory for Experimental Immunology, Ghent University, Belgium
Katrien De Mulder
Laboratory for Experimental Immunology, Ghent University, Belgium
Yasmine Van Caeneghem
Laboratory for Experimental Immunology, Ghent University, Belgium
Greet Verstichel
Laboratory for Experimental Immunology, Ghent University, Belgium
Nadine Van Roy
Center for Medical Genetics, Ghent University, Belgium
Björn Menten
Center for Medical Genetics, Ghent University, Belgium
Imke Velghe
Laboratory for Experimental Immunology, Ghent University, Belgium
Jan Philippé
Department of Clinical Biology, Microbiology and Immunology, Ghent University Hospital, Belgium
Dominique De Bleser
Red Cross Flanders, Ghent, Belgium
Bart N. Lambrecht
Laboratory of Immunoregulation and Mucosal Immunology, Department of Pulmonary Medicine, Ghent University Hospital, Belgium;Flanders Institute for Biotechnology (VIB) Inflammation Research Center, Ghent University, Belgium
Tom Taghon
Laboratory for Experimental Immunology, Ghent University, Belgium
Georges Leclercq
Laboratory for Experimental Immunology, Ghent University, Belgium
Tessa Kerre
Laboratory for Experimental Immunology, Ghent University, Belgium
Bart Vandekerckhove
Laboratory for Experimental Immunology, Ghent University, Belgium
Although hematopoietic precursor activity can be generated in vitro from human embryonic stem cells, there is no solid evidence for the appearance of multipotent, self-renewing and transplantable hematopoietic stem cells. This could be due to short half-life of hematopoietic stem cells in culture or, alternatively, human embryonic stem cell-initiated hematopoiesis may be hematopoietic stem cell-independent, similar to yolk sac hematopoiesis, generating multipotent progenitors with limited expansion capacity. Since a MYB was reported to be an excellent marker for hematopoietic stem cell-dependent hematopoiesis, we generated a MYB-eGFP reporter human embryonic stem cell line to study formation of hematopoietic progenitor cells in vitro. We found CD34+ hemogenic endothelial cells rounding up and developing into CD43+ hematopoietic cells without expression of MYB-eGFP. MYB-eGFP+ cells appeared relatively late in embryoid body cultures as CD34+CD43+CD45−/lo cells. These MYB-eGFP+ cells were CD33 positive, proliferated in IL-3 containing media and hematopoietic differentiation was restricted to the granulocytic lineage. In agreement with data obtained on murine Myb−/− embryonic stem cells, bright eGFP expression was observed in a subpopulation of cells, during directed myeloid differentiation, which again belonged to the granulocytic lineage. In contrast, CD14+ macrophage cells were consistently eGFP− and were derived from eGFP-precursors only. In summary, no evidence was obtained for in vitro generation of MYB+ hematopoietic stem cells during embryoid body cultures. The observed MYB expression appeared late in culture and was confined to the granulocytic lineage.
