title: Regulation of primitive hematopoiesis in zebrafish embryos by the death receptor gene.
abstract: OBJECTIVE
  We investigated the regulatory mechanism of primitive hematopoiesis in zebrafish (Danio rerio) embryos with particular reference to the role of a death receptor (zDR) gene, based on a morpholino (MO) knockdown approach.
  
  
  METHODS
  MOs targeting the zDR and chordin (Chd) were injected into naturally spawned embryos at one- to four-cell stage. A random sequence (RS) MO was used as a control. Effects on hemoglobin formation (Hb), apoptosis, and lineage-specific gene expression were examined. Embryos injected with zDR, Chd, and RS-MOs were denoted zDR(mo), zChd(mo), and zRS(mo), respectively. Those co-injected with Chd+zDR-MOs and Chd+RS-MOs were abbreviated zChd+DR(mo) and zChd+RS(mo).
  
  
  RESULTS
  zDR mRNA expression was restricted to the intermediate cell mass of wild-type (WT) and zChd(mo) embryos. At 48 hours postfertilization, zDR(mo) embryos showed increased Hb compared with WT or zRS(mo) embryos (2.36 x 10(-2) +/- 1.13 x 10(-3) vs 1.85 x 10(-2) +/- 5.60 x 10(-4) vs 1.79 x 10(-2) +/- 1.31 x 10(-3) U, p < 0.05). zChd+DR(mo) embryos also showed increased Hb compared with zChd(mo) or zChd+RS(mo) embryos (4.60 x 10(-2) +/- 2.79 x 10(-3) vs 3.17 x 10(-2) +/- 1.07 x 10(-3) vs 3.05 x 10(-2) +/- 1.25 x 10(-3) U, p < 0.05). zDR-MO reduced apoptosis, as shown by reduced terminal transferase-mediated dUTP nick end-labeling staining in zChd+DR(mo) compared with zChd+RS(mo) embryos and caspase-3 activity in zDR(mo) vs zRS(mo) (0.525 +/- 0.094 vs 0.953 +/- 0.113 U, p < 0.05), and zChd+DR(mo) vs zChd+RS(mo) embryos (0.247 +/- 0.121 vs 1.180 +/- 0.082, p < 0.05). zChd+DR(mo) embryos showed upregulation of erythroid-specific embryonic hemoglobin gene expression but not that of a myeloid-specific myeloperoxidase gene.
  
  
  CONCLUSIONS
  Knockdown of zDR in zebrafish embryos decreased apoptosis and increased Hb, suggesting that zDR may regulate primitive hematopoiesis during development.
authors.affiliation: Department of Medicine, University of Hong Kong, Hong Kong.
authors.forename: Tommy T
authors.initials: TT
authors.lastname: Kwan
authors.forename: Raymond
authors.initials: R
authors.lastname: Liang
authors.forename: Catherine M
authors.initials: CM
authors.lastname: Verfaillie
authors.forename: Stephen C
authors.initials: SC
authors.lastname: Ekker
authors.forename: Li C
authors.initials: LC
authors.lastname: Chan
authors.forename: Shuo
authors.initials: S
authors.lastname: Lin
authors.forename: Anskar Y H
authors.initials: AY
authors.lastname: Leung
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3; D020169:Caspases / Q000187:drug effects / Q000378:metabolism; D004195:Disease Models, Animal; D004625:Embryo, Nonmammalian; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D006023:Glycoproteins / Q000008:administration & dosage; D006410:Hematopoiesis / Q000187:drug effects / Q000235:genetics / Q000502:physiology*; D006454:Hemoglobins / Q000096:biosynthesis / Q000187:drug effects; D036341:Intercellular Signaling Peptides and Proteins / Q000008:administration & dosage; D009025:Morpholines / Q000008:administration & dosage; D018124:Receptors, Tumor Necrosis Factor / Q000008:administration & dosage* / Q000235:genetics* / Q000378:metabolism; D015027:Zebrafish / Q000196:embryology* / Q000235:genetics / Q000378:metabolism*
chemical_list: D006023:Glycoproteins; D006454:Hemoglobins; D036341:Intercellular Signaling Peptides and Proteins; D009025:Morpholines; D018124:Receptors, Tumor Necrosis Factor; C414429:hematopoietic death receptor; C054234:chordin; D053148:Caspase 3; D020169:Caspases