title: Effect of the oxidative phosphorylation uncoupler 2,4-dinitrophenol on hypoxia-inducible factor-regulated gene expression in bovine blastocysts.
abstract: In cattle embryos, development to the blastocyst stage is improved in the presence of 10 micro;m 2,4-dinitrophenol (DNP), an uncoupler of oxidative phosphorylation, coincident with an increase in glycolytic activity following embryonic genome activation. The present study examined redox-sensitive gene expression and embryo development in response to the addition of DNP post-compaction. 2,4-Dinitrophenol increased the expression of hypoxia-inducible factor 1alpha and 2alpha (HIF1alpha, HIF2alpha) mRNA. Although HIF1alpha protein remained undetectable in bovine blastocysts, HIF2alpha protein was localised within the nucleus of trophectoderm and inner cell mass (ICM) cells of blastocysts cultured in the presence or absence of DNP, with a slight increase in staining evident within the ICM in blastocysts cultured in the presence of DNP. However, the expression of GLUT1 and VEGF mRNA, genes known to be regulated by HIFs, was unaffected by the addition of DNP to the culture. Although the development of Grade 1 and 2 blastocysts was unaltered by the addition of DNP post compaction in the present study, a significant increase in the proportion of ICM cells was observed. Results indicate that 10 microm DNP improves the quality of bovine embryos, coincident with increased HIF2alpha protein localisation within ICM cells and increased HIFalpha mRNA levels. Therefore, the results demonstrate redox-regulated expression of HIF2.
authors.affiliation: Research Centre for Reproductive Health, Department of Obstetrics and Gynaecology, University of Adelaide, Queen Elizabeth Hospital, Woodville, SA, Australia. alexandra.harvey@adelaide.edu.au
authors.forename: A J
authors.initials: AJ
authors.lastname: Harvey
authors.forename: K L
authors.initials: KL
authors.lastname: Kind
authors.forename: J G
authors.initials: JG
authors.lastname: Thompson
mesh_terms: D019297:2,4-Dinitrophenol / Q000494:pharmacology*; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000096:biosynthesis / Q000235:genetics*; D001755:Blastocyst / Q000187:drug effects* / Q000378:metabolism; D002417:Cattle; D046149:Embryo Culture Techniques; D005786:Gene Expression Regulation / Q000187:drug effects*; D051272:Glucose Transporter Type 1 / Q000235:genetics; D051795:Hypoxia-Inducible Factor 1, alpha Subunit / Q000096:biosynthesis / Q000235:genetics*; D010084:Oxidation-Reduction; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014475:Uncoupling Agents / Q000494:pharmacology*; D042461:Vascular Endothelial Growth Factor A / Q000235:genetics; D000099162:Endothelial PAS Domain-Containing Protein 1
chemical_list: D019297:2,4-Dinitrophenol; D051792:Basic Helix-Loop-Helix Proteins; D051272:Glucose Transporter Type 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D012333:RNA, Messenger; D014475:Uncoupling Agents; D042461:Vascular Endothelial Growth Factor A; D000099162:Endothelial PAS Domain-Containing Protein 1