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First published online August 3, 2006
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2006-0199v1
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Submitted on April 7, 2006
Accepted on July 26, 2006

Embryonic Stem Cells

Sequential analysis of {alpha}- and {beta}-globin gene expression during erythropoietic differentiation from primate embryonic stem cells

Katsutsugu Umeda 1, Toshio Heike 1, Mami Nakata-Hizume 1, Akira Niwa 1, Masato Arai 1, Gen Shinoda 1, Feng Ma 1, Hirofumi Suemori 2, Hong Yuan Luo 3, David H. K. Chui 3, Ryuzo Torii 4, Masabumi Shibuya 5, Norio Nakatsuji 6, Tatsutoshi Nakahata 1*

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 Department of Medicine, Boston University School of Medicine, Boston, Massachusetts
4 Research Center for Animal Life Science, Shiga University of Medical Sciences, Shiga, Japan
5 Division of Genetics, Institute of Medical Sciences, University of Tokyo, Tokyo, Japan
6 Department of Development and Differentiation, Institute for Frontier Medical Sciences, Kyoto University, Kyoto, Japan

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


  Abstract

The temporal pattern of embryonic, fetal, and adult globin expression in the {alpha} ({zeta} -> {alpha}), and {beta} ({epsilon} -> {gamma} and {gamma} -> {beta}) cluster were quantitatively analyzed at the transcriptional and translational levels in erythrocytes induced from primate embryonic stem cells in vitro. When VEGF receptor-2high CD34+ cells were harvested and reseeded onto OP9 stromal cells, two-wave erythropoiesis occurred sequentially. Immunostaining and real-time RT-PCR analyses of floating mature erythrocytes revealed that globin switches occurred in parallel with the erythropoietic transition. Colony-forming assays showed replacement of primitive clonogenic progenitor cells with definitive cells during culturing. A decline in embryonic {zeta}- and {epsilon}-globin expression at the translational level occurred in individual definitive erythroid progenitors. Expression of {beta}-globin in individual definitive erythroid progenitors was up-regulated in the presence of OP9 stromal cells. Thus, this system reproduces early hematopoietic development in vitro and can serve as a model for analyzing the mechanisms of the globin switch in humans.

Key Words. ES cells, erythroid progenitors




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 Proc. Natl. Acad. Sci. USAHome page
F. Ma, Y. Ebihara, K. Umeda, H. Sakai, S. Hanada, H. Zhang, Y. Zaike, E. Tsuchida, T. Nakahata, H. Nakauchi, et al.
Generation of functional erythrocytes from human embryonic stem cell-derived definitive hematopoiesis
PNAS, September 2, 2008; 105(35): 13087 - 13092.
[Abstract] [Full Text] [PDF]


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