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Prepublished online as a Blood First Edition Paper on February 20, 2003; DOI 10.1182/blood-2002-09-2799.
Submitted September 30, 2002
Accepted January 21, 2003
Erythropoiesis from acetyl LDL incorporating endothelial cells at the pre-liver stage
Daisuke Sugiyama*, Minetaro Ogawa, Imiko Hirose, Thierry Jaffredo, Ken-ichi Arai, and Kohichiro Tsuji
Division of Molecular and Developmental Biology, Department of Basic Medical Science, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan
Division of Cellular Therapy, Advanced Clinical Research Center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan
Department of Cell Differentiation, Institute of Molecular Embryology and Genetics, Kumamoto University, Kumamoto, Japan
Developmental Biology Unit, University Pierre and Marie Curie, Paris, France
* Corresponding author; email: Daisuke.Sugiyama{at}snv.jussieu.fr.
Erythropoiesis is characterized by two distinct waves of production during mouse embryogenesis: a primitive one originating from the yolk sac (YS) and a definitive one produced from both the YS and the embryo proper. How this last wave is generated remains unclear. We put forward the hypothesis that erythroid cells could be generated by endothelial cells (ECs). To investigate this problem in depth, we have designed a method to label ECs at 10 days post-coitum. This label associates two techniques: an intracardiac inoculation that allows molecules to be delivered into the blood stream, followed by a whole embryo culture period. DiI-conjugated acetylated low-density lipoproteins (Ac-LDL-DiI) was used to specifically tag ECs from the inside. One hour after inoculation, DiI staining was found along the entire endothelial tree. FACS analysis revealed that DiI+ cells were CD31+, CD34+ and CD45-, an antigen make-up characteristic for the endothelial lineage. Twelve hours after inoculation, 43% of DiI+ circulating cells belonged to the erythroid lineage. These cells expressed Ter 119 and displayed an adult globin chain arrangement, thus belonged to the definitive lineage as confirmed in erythroid colony formation. The rest of the cells likely represent committed white blood cells or multi-potent progenitors as revealed by a mix-colony formation. Beyond the 29-somite stage, the proportion of the DiI+ erythroid cells gradually decreased. These results demonstrate, for the first time in the mouse embryo, the generation of hematopoietic cells from an endothelial intermediate, using an in vivo tracing. We thus provide evidence for a release of these cells into the circulation and put forward the hypothesis that these cells are able to colonize the fetal liver and generate definitive erythrocytes in vivo.
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