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Title: Identification of a population of cells with hematopoietic stem cell properties in mouse aorta-gonad-mesonephros cultures

Abstract

The aorta-gonad-mesonephros (AGM) region is a primary source of definitive hematopoietic cells in the midgestation mouse embryo. In cultures of dispersed AGM regions, adherent cells containing endothelial cells are observed first, and then non-adherent hematopoietic cells are produced. Here we report on the characterization of hematopoietic cells that emerge in the AGM culture. Based on the expression profiles of CD45 and c-Kit, we defined three cell populations: CD45{sup low} c-Kit{sup +} cells that had the ability to form hematopoietic cell colonies in methylcellulose media and in co-cultures with stromal cells; CD45{sup low} c-Kit{sup -} cells that showed a granulocyte morphology; CD45{sup high} c-Kit{sup low/-} that exhibited a macrophage morphology. In co-cultures of OP9 stromal cells and freshly prepared AGM cultures, CD45{sup low} c-Kit{sup +} cells from the AGM culture had the abilities to reproduce CD45{sup low} c-Kit{sup +} cells and differentiate into CD45{sup low} c-Kit{sup -} and CD45{sup high} c-Kit{sup low/-} cells, whereas CD45{sup low} c-Kit{sup -} and CD45{sup high} c-Kit{sup low/-} did not produce CD45{sup low} c-Kit{sup +} cells. Furthermore, CD45{sup low} c-Kit{sup +} cells displayed a long-term repopulating activity in adult hematopoietic tissue when transplanted into the liver of irradiated newborn mice. These results indicate that CD45{sup low}more » c-Kit{sup +} cells from the AGM culture have the potential to reconstitute multi-lineage hematopoietic cells.« less

Authors:
 [1];  [1];  [2];  [3];  [4]
  1. Division of Cell Fate Modulation, Institute of Molecular Embryology and Genetics, Kumamoto University, 2-2-1 Honjo, Kumamoto (Japan)
  2. Division of Hematopoiesis, Center for AIDS Research, Kumamoto University, 2-2-1 Honjo, Kumamoto (Japan)
  3. Center for Animal Resources and Development, Kumamoto University, 2-2-1 Honjo, Kumamoto (Japan)
  4. Division of Cell Fate Modulation, Institute of Molecular Embryology and Genetics, Kumamoto University, 2-2-1 Honjo, Kumamoto (Japan). E-mail: taga@kaiju.medic.kumamoto-u.ac.jp
Publication Date:
OSTI Identifier:
20972129
Resource Type:
Journal Article
Resource Relation:
Journal Name: Experimental Cell Research; Journal Volume: 313; Journal Issue: 5; Other Information: DOI: 10.1016/j.yexcr.2006.12.024; PII: S0014-4827(06)00517-9; Copyright (c) 2007 Elsevier Science B.V., Amsterdam, The Netherlands, All rights reserved; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United States
Language:
English
Subject:
60 APPLIED LIFE SCIENCES; AORTA; BLOOD FORMATION; EMBRYOS; GONADS; INFANTS; IRRADIATION; LEUKOCYTES; LIVER; MACROPHAGES; MICE; MORPHOLOGY; STEM CELLS

Citation Formats

Nobuhisa, Ikuo, Ohtsu, Naoki, Okada, Seiji, Nakagata, Naomi, and Taga, Tetsuya. Identification of a population of cells with hematopoietic stem cell properties in mouse aorta-gonad-mesonephros cultures. United States: N. p., 2007. Web. doi:10.1016/j.yexcr.2006.12.024.
Nobuhisa, Ikuo, Ohtsu, Naoki, Okada, Seiji, Nakagata, Naomi, & Taga, Tetsuya. Identification of a population of cells with hematopoietic stem cell properties in mouse aorta-gonad-mesonephros cultures. United States. doi:10.1016/j.yexcr.2006.12.024.
Nobuhisa, Ikuo, Ohtsu, Naoki, Okada, Seiji, Nakagata, Naomi, and Taga, Tetsuya. Sat . "Identification of a population of cells with hematopoietic stem cell properties in mouse aorta-gonad-mesonephros cultures". United States. doi:10.1016/j.yexcr.2006.12.024.
@article{osti_20972129,
title = {Identification of a population of cells with hematopoietic stem cell properties in mouse aorta-gonad-mesonephros cultures},
author = {Nobuhisa, Ikuo and Ohtsu, Naoki and Okada, Seiji and Nakagata, Naomi and Taga, Tetsuya},
abstractNote = {The aorta-gonad-mesonephros (AGM) region is a primary source of definitive hematopoietic cells in the midgestation mouse embryo. In cultures of dispersed AGM regions, adherent cells containing endothelial cells are observed first, and then non-adherent hematopoietic cells are produced. Here we report on the characterization of hematopoietic cells that emerge in the AGM culture. Based on the expression profiles of CD45 and c-Kit, we defined three cell populations: CD45{sup low} c-Kit{sup +} cells that had the ability to form hematopoietic cell colonies in methylcellulose media and in co-cultures with stromal cells; CD45{sup low} c-Kit{sup -} cells that showed a granulocyte morphology; CD45{sup high} c-Kit{sup low/-} that exhibited a macrophage morphology. In co-cultures of OP9 stromal cells and freshly prepared AGM cultures, CD45{sup low} c-Kit{sup +} cells from the AGM culture had the abilities to reproduce CD45{sup low} c-Kit{sup +} cells and differentiate into CD45{sup low} c-Kit{sup -} and CD45{sup high} c-Kit{sup low/-} cells, whereas CD45{sup low} c-Kit{sup -} and CD45{sup high} c-Kit{sup low/-} did not produce CD45{sup low} c-Kit{sup +} cells. Furthermore, CD45{sup low} c-Kit{sup +} cells displayed a long-term repopulating activity in adult hematopoietic tissue when transplanted into the liver of irradiated newborn mice. These results indicate that CD45{sup low} c-Kit{sup +} cells from the AGM culture have the potential to reconstitute multi-lineage hematopoietic cells.},
doi = {10.1016/j.yexcr.2006.12.024},
journal = {Experimental Cell Research},
number = 5,
volume = 313,
place = {United States},
year = {Sat Mar 10 00:00:00 EST 2007},
month = {Sat Mar 10 00:00:00 EST 2007}
}
  • Long-term reconstituting hematopoietic stem cells first arise from the aorta of the aorta-gonad-mesonephros (AGM) region in a mouse embryo. We have previously reported that in cultures of the dispersed AGM region, CD45{sup low}c-Kit{sup +} cells possess the ability to reconstitute multilineage hematopoietic cells, but investigations are needed to show that this is not a cultured artifact and to clarify when and how this population is present. Based on the expression profile of CD45 and c-Kit in freshly dissociated AGM cells from embryonic day 9.5 (E9.5) to E12.5 and aorta cells in the AGM from E13.5 to E15.5, we defined sixmore » cell populations (CD45{sup -}c-Kit{sup -}, CD45{sup -}c-Kit{sup low}, CD45{sup -}c-Kit{sup high}, CD45{sup low}c-Kit{sup high}, CD45{sup high}c-Kit{sup high}, and CD45{sup high}c-Kit{sup very} {sup low}). Among these six populations, CD45{sup low}c-Kit{sup high} cells were most able to form hematopoietic cell colonies, but their ability decreased after E11.5 and was undetectable at E13.5 and later. The CD45{sup low}c-Kit{sup high} cells showed multipotency in vitro. We demonstrated further enrichment of hematopoietic activity in the Hoechst dye-effluxing side population among the CD45{sup low}c-Kit{sup high} cells. Here, we determined that CD45{sup low}c-Kit{sup high} cells arise from the lateral plate mesoderm using embryonic stem cell-derived differentiation system. In conclusion, CD45{sup low}c-Kit{sup high} cells are the major hematopoietic cells of mouse AGM.« less
  • We investigated whether the in vitro differentiation of ES cells into haematopoietic progenitors could be enhanced by exposure to the aorta-gonadal-mesonephros (AGM) microenvironment that is involved in the generation of haematopoietic stem cells (HSC) during embryonic development. We established a co-culture system that combines the requirements for primary organ culture and differentiating ES cells and showed that exposure of differentiating ES cells to the primary AGM region results in a significant increase in the number of ES-derived haematopoietic progenitors. Co-culture of ES cells on the AM20-1B4 stromal cell line derived from the AGM region also increases haematopoietic activity. We concludemore » that factors promoting the haematopoietic activity of differentiating ES cells present in primary AGM explants are partially retained in the AM20.1B4 stromal cell line and that these factors are likely to be different to those required for adult HSC maintenance.« less
  • The aorta-gonad-mesonephros (AGM) region is involved in the generation and maintenance of the first definitive hematopoietic stem cells (HSCs). A mouse AGM-derived cell line, AGM-S3, was shown to support the development of HSCs. To elucidate the molecular mechanisms regulating early hematopoiesis, we obtained subclones from AGM-S3, one of which was hematopoiesis supportive (S3-A9) and the other one of which was non-supportive (S3-A7), and we analyzed their gene expression profiles by gene chip analysis. In the present study, we found that Glypican-1 (GPC1) was highly expressed in the supportive subclone AGM-S3-A9. Over-expression of GPC1 in non-supportive cells led to the proliferationmore » of progenitor cells in human cord blood when cocultured with the transfected-stromal cells. Thus, GPC1 may have an important role in the establishment of a microenvironment that supports early events in hematopoiesis.« less
  • The authors study changes in the number of splenic exocolonies and also postradiation repopulation of hematopoietic organs in recipients after receiving an injection of bone marrow treated with mouse antibrain serum (MABS), with or without the addition of thymocytes in the later stages. The recipient mice were irradiated with /sup 60/Co gamma rays in a dose of 8.5-8.0 Gy, on the Luch-1 radiotherapeutic apparatus 18 h before receiving the injection of bone marrow and thymocytes. It is postulated that MABS inactivates a population of cells in the bone marrow which are invloved in the control of colony-forming-units (CFUs) proliferation. Thismore » population is restored in the animal by the 14th day after irradiation and of injection of MABS-treated bone marrow.« less