Hematopoietic Stem Cell Formation in Zebrafish Is Regulated in a Temporally Distinct Manner by Estrogen Receptor 2

• Published in: Blood 2011-11, Vol.118 (21), p.1268-1268
• Main Authors: Carroll, Kelli J., Dovey, Michael C, Cutting, Claire C, Sheward, Lea Vedder, Harris, James M, Goessling, Wolfram, North, Trista E.
• Format: Article
• Language: English
• ISSN: 0006-4971; 1528-0020
• DOI: 10.1182/blood.V118.21.1268.1268

Abstract 1268 The intrinsic signaling pathways regulating hematopoietic stem cells (HSC) are increasingly understood; in contrast, less is known about the potential effect of exposure to environmental factors, such as xenoestrogens, on the formation of HSCs. RUNX1 (AML1) is a highly conserved transcription factor that is required for definitive HSC induction and is also the target of many chromosomal alterations in leukemia. Through a chemical genetic screen, estrogen-related compounds were identified as modifiers of runx1 expression in the zebrafish. Exposure to 17b-estradiol (E2) throughout hematopoietic development (5 somites (som) to 36 hours post fertilization (hpf)) significantly decreased the number of runx1+ HSCs in the zebrafish Aorta-Gonad-Mesonephros Region (AGM) compared to controls (n≥25–50 embryos /condition). Use of the nonspecific estrogen receptor inhibitor fulvestrant confirmed that estrogen was required for HSC regulation and functioned through classical estrogen receptors. Microarray analysis of FACS-sorted cell populations during zebrafish development demonstrated differential spatio-temporal regulation and expression of esr1 (esrα) and esr2a/b (esrβ) in vascular and hematopoietic cell types; use of an ERE-GFP reporter fish verified that estrogen signaling is active during this stage of embryonic development. During the primitive wave of hematopoiesis, exposure to E2 and the esr1-agonist PPT significantly enhanced red blood cell number as seen by in situ hybridization for embryonic globin (hbbe3) and quantified by fluorescent microscopy and FACS analysis of the Tg(globin:GFP) line. Conversely, the esr2-specific agonist DPN diminished definitive HSC formation after exposure from 5 som to 24 hpf; this phenotype was mediated by disruption of vessel formation, as indicated by flk1 (kdrl) expression, and alteration in the assignment of artery-vein identity. Alterations in arterial specification appear to be mediated by the Notch/VEGF pathway. E2 exposure from 5 somites to 36 hpf decreased GFP expression in notch reporter fish as well as expression of deltaC and notch5 by in situ hybridization. Interestingly, when exposure to E2 or DPN occurred from 24 – 36 hpf, after arterial establishment and initiation of blood flow, estrogen treatment enhanced HSC formation; this was confirmed by FACS analysis and fluorescent microscopy using the Tg(cmyb:eGFP) and Tg(-6.0itga2b:eGFP)la2 (CD41:GFP) HSC reporter lines. E2 treatment was found to elicit both