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Sci Rep. 2019 Oct 2;9(1):14205. doi: 10.1038/s41598-019-50672-5.

Enrichment of hematopoietic stem/progenitor cells in the zebrafish kidney.

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Faculty of Biological Science and Technology, Institute of Science and Engineering, Kanazawa University, Kanazawa, Ishikawa, Japan.
Division of Life Sciences, Graduate School of Natural Science and Technology, Kanazawa University, Kanazawa, Ishikawa, Japan.
Department of Life Science, Medical Research Institute, Kanazawa Medical University, Uchinada, Ishikawa, Japan.
Department of Applied Biological Science, Faculty of Science and Technology, Tokyo University of Science, Noda, Chiba, Japan.
Laboratory of Comparative Immunology, Department of Veterinary Medicine, Nihon University, Fujisawa, Kanagawa, Japan.
Department of Cellular and Molecular Medicine, University of California at San Diego, La Jolla, CA, USA.


Hematopoietic stem cells (HSCs) maintain the entire blood system throughout life and are utilized in therapeutic approaches for blood diseases. Prospective isolation of highly purified HSCs is crucial to understand the molecular mechanisms underlying regulation of HSCs. The zebrafish is an elegant genetic model for the study of hematopoiesis due to its many unique advantages. It has not yet been possible, however, to purify HSCs in adult zebrafish due to a lack of specific HSC markers. Here we show the enrichment of zebrafish HSCs by a combination of two HSC-related transgenes, gata2a:GFP and runx1:mCherry. The double-positive fraction of gata2a:GFP and runx1:mCherry (gata2a+ runx1+) was detected at approximately 0.16% in the kidney, the main hematopoietic organ in teleosts. Transcriptome analysis revealed that gata2a+ runx1+ cells showed typical molecular signatures of HSCs, including upregulation of gata2b, gfi1aa, runx1t1, pbx1b, and meis1b. Transplantation assays demonstrated that long-term repopulating HSCs were highly enriched within the gata2a+ runx1+ fraction. In contrast, colony-forming assays showed that gata2a- runx1+ cells abundantly contain erythroid- and/or myeloid-primed progenitors. Thus, our purification method of HSCs in the zebrafish kidney is useful to identify molecular cues needed to regulate self-renewal and differentiation of HSCs.

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