Hemogenic and nonhemogenic endothelium can be distinguished by the activity of fetal liver kinase (Flk)-1 promoter/enhancer during mouse embryogenesis.
Accumulating evidence in various species has suggested that the origin of definitive hematopoiesis is associated with a special subset of endothelial cells (ECs) that maintain the potential to give rise to hematopoietic cells (HPCs). In this study, we demonstrated that a combination of 5'-flanking region and 3' portion of the first intron of the Flk-1 gene (Flk-1 p/e) that has been implicated in endothelium-specific gene expression distinguishes prospectively the EC that has lost hemogenic activity. We assessed the activity of this Flk-1 p/e by embryonic stem (ES) cell differentiation culture and transgenic mice by using the GFP gene conjugated to this unit. The expression of GFP differed from that of the endogenous Flk-1 gene in that it is active in undifferentiated ES cells and inactive in Flk-1(+) lateral mesoderm. Flk-1 p/e becomes active after generation of vascular endothelial (VE)-cadherin(+) ECs. Emergence of GFP(-) ECs preceded that of GFP(+) ECs, and, finally, most ECs expressed GFP both in vitro and in vivo. Cell sorting experiments demonstrated that only GFP(-) ECs could give rise to HPCs and preferentially expressed Runx1 and c-Myb genes that are required for the definitive hematopoiesis. Integration of both GFP(+) and GFP(-) ECs was observed in the dorsal aorta, but cell clusters appeared associated only to GFP(-) ECs. These results indicate that activation of Flk-1 p/e is associated with a process that excludes HPC potential from the EC differentiation pathway and will be useful for investigating molecular mechanisms underlying the divergence of endothelial and hematopoietic lineages.
Hirai
Hideyo
H
Department of Microbiology, Kyoto Prefectural University of Medicine, Kyoto, Japan. hhirai@basic.kpu-m.ac.jp
Ogawa
Minetaro
M
Suzuki
Norio
N
Yamamoto
Masayuki
M
Breier
Georg
G
Mazda
Osam
O
Imanishi
Jiro
J
Nishikawa
Shin-Ichi
S
D000818:Animals; D018929:Cell Culture Techniques; D019070:Cell Lineage; D004622:Embryo, Mammalian / Q000166:cytology*; D004730:Endothelium, Vascular / Q000166:cytology*; D049452:Green Fluorescent Proteins; D006410:Hematopoiesis; D006412:Hematopoietic Stem Cells / Q000166:cytology; D008164:Luminescent Proteins / Q000096:biosynthesis / Q000235:genetics; D051379:Mice; D011401:Promoter Regions, Genetic / Q000502:physiology*; D013234:Stem Cells / Q000166:cytology*; D014157:Transcription Factors / Q000032:analysis; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000235:genetics*
D008164:Luminescent Proteins; D014157:Transcription Factors; D049452:Green Fluorescent Proteins; D040301:Vascular Endothelial Growth Factor Receptor-2