Identification and characterization of hemoangiogenic progenitors during cynomolgus monkey embryonic stem cell differentiation

¹ Department of Pediatrics, Graduate School of Medicine, Kyoto University, Kyoto, Japan
² Laboratory of Embryonic Stem Cell Research, Stem Cell Research Center, Institute for Frontier Medical Sciences, Kyoto University, Kyoto, Japan
³ Deparment of Medicine, Boston University School of Medicine, Boston, Massachusetts
⁴ Research Center for Animal Life Science, Shiga University of Medical Science, Shiga, Japan
⁵ Division of Genetics, Institute of Medical Science, University of Tokyo, Tokyo, Japan
⁶ Department of Development and Differentiation, Institute for Frontier Medical Science, Kyoto University, Kyoto, Japan

^* To whom correspondence should be addressed. E-mail: tnakaha{at}kuhp.kyoto-u.ac.jp.

	Abstract

We identified intermediate-stage progenitor cells that have the potential to differentiate into hematopoietic and endothelial lineages from nonhuman primate embryonic stem (ES) cells. Sequential fluorescence activated cell sorting (FACS) and immunostaining analyses showed that when ES cells were cultured in an OP9 coculture system, both lineages developed after the emergence of two hemoangiogenic progenitor-bearing cell fractions, namely, vascular endothelial growth factor receptor (VEGFR)-2^high CD34^- and VEGFR-2^high CD34⁺ cells. Exogenous VEGF increased the proportion of VEGFR-2^high cells, particularly that of VEGFR-2^high CD34⁺ cells, in a dose-dependent manner. While either population of VEGFR-2^high cells could differentiate into primitive and definitive hematopoietic cells (HCs), as well as endothelial cells (ECs), the VEGFR-2^high CD34⁺ cells had greater hemoangiogenic potential. Both lineages developed from VEGFR-2^high CD34^- or VEGFR-2^high CD34⁺ precursor at the single cell level, which strongly supports the existence of hemangioblasts in these cell fractions. Thus, this culture system allows differentiation into the HC and EC lineages to be defined by surface markers. These observations should facilitate further studies both on early developmental processes and on regeneration therapies in human.

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