title: A mRNA landscape of bovine embryos after standard and MAPK-inhibited culture conditions: a comparative analysis.
abstract: BACKGROUND
  Genes and signalling pathways involved in pluripotency have been studied extensively in mouse and human pre-implantation embryos and embryonic stem (ES) cells. The unsuccessful attempts to generate ES cell lines from other species including cattle suggests that other genes and pathways are involved in maintaining pluripotency in these species. To investigate which genes are involved in bovine pluripotency, expression profiles were generated from morula, blastocyst, trophectoderm and inner cell mass (ICM) samples using microarray analysis. As MAPK inhibition can increase the NANOG/GATA6 ratio in the inner cell mass, additionally blastocysts were cultured in the presence of a MAPK inhibitor and changes in gene expression in the inner cell mass were analysed.
  
  
  RESULTS
  Between morula and blastocyst 3,774 genes were differentially expressed and the largest differences were found in blastocyst up-regulated genes. Gene ontology (GO) analysis shows lipid metabolic process as the term most enriched with genes expressed at higher levels in blastocysts. Genes with higher expression levels in morulae were enriched in the RNA processing GO term. Of the 497 differentially expressed genes comparing ICM and TE, the expression of NANOG, SOX2 and POU5F1 was increased in the ICM confirming their evolutionary preserved role in pluripotency. Several genes implicated to be involved in differentiation or fate determination were also expressed at higher levels in the ICM. Genes expressed at higher levels in the ICM were enriched in the RNA splicing and regulation of gene expression GO term. Although NANOG expression was elevated upon MAPK inhibition, SOX2 and POU5F1 expression showed little increase. Expression of other genes in the MAPK pathway including DUSP4 and SPRY4, or influenced by MAPK inhibition such as IFNT, was down-regulated.
  
  
  CONCLUSIONS
  The data obtained from the microarray studies provide further insight in gene expression during bovine embryonic development. They show an expression profile in pluripotent cells that indicates a pluripotent, epiblast-like state. The inability to culture ICM cells as stem cells in the presence of an inhibitor of MAPK activity together with the reported data indicates that MAPK inhibition alone is not sufficient to maintain a pluripotent character in bovine cells.
authors.affiliation: Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, Utrecht, 3584 CM, The Netherlands. b.brinkhof@uu.nl.
authors.forename: Bas
authors.initials: B
authors.lastname: Brinkhof
authors.affiliation: Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, Utrecht, 3584 CM, The Netherlands. H.T.A.vanTol@uu.nl.
authors.forename: Helena T A
authors.initials: HT
authors.lastname: van Tol
authors.affiliation: University Medical Center Utrecht, Molecular Cancer Research, PO Box 85060, Utrecht, 3508 AB, The Netherlands. M.J.A.GrootKoerkamp@umcutrecht.nl.
authors.forename: Marian J A
authors.initials: MJ
authors.lastname: Groot Koerkamp
authors.affiliation: Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, University Utrecht, Yalelaan 108, Utrecht, 3584 CM, The Netherlands. F.M.Riemers@uu.nl.
authors.forename: Frank M
authors.initials: FM
authors.lastname: Riemers
authors.affiliation: Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, Utrecht, 3584 CM, The Netherlands. sascha_ijzer@hotmail.com.
authors.forename: Sascha G
authors.initials: SG
authors.lastname: IJzer
authors.affiliation: Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, Utrecht, 3584 CM, The Netherlands. kavehm@kth.se.|BioTalentum Ltd, Aulich L u.26, Gödöllő, 2100, Hungary. kavehm@kth.se.
authors.forename: Kaveh
authors.initials: K
authors.lastname: Mashayekhi
authors.affiliation: Department of Infectious Diseases and Immunology, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 1, Utrecht, 3584 CL, The Netherlands. H.P.Haagsman@uu.nl.
authors.forename: Henk P
authors.initials: HP
authors.lastname: Haagsman
authors.affiliation: Department of Farm Animal Health, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 104, Utrecht, 3584 CM, The Netherlands. B.A.J.Roelen@uu.nl.
authors.forename: Bernard A J
authors.initials: BA
authors.lastname: Roelen
mesh_terms: D000818:Animals; D001549:Benzamides / Q000494:pharmacology; D053624:Blastocyst Inner Cell Mass / Q000187:drug effects / Q000378:metabolism*; D002417:Cattle / Q000196:embryology* / Q000235:genetics / Q000378:metabolism; D002478:Cells, Cultured; D004159:Diphenylamine / Q000031:analogs & derivatives / Q000494:pharmacology; D046149:Embryo Culture Techniques; D018507:Gene Expression Regulation, Developmental / Q000187:drug effects; D020929:Mitogen-Activated Protein Kinase Kinases / Q000378:metabolism*; D009028:Morula / Q000187:drug effects / Q000378:metabolism*; D039904:Pluripotent Stem Cells / Q000378:metabolism; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism*
chemical_list: D001549:Benzamides; D047428:Protein Kinase Inhibitors; D012333:RNA, Messenger; C506614:mirdametinib; D004159:Diphenylamine; D020929:Mitogen-Activated Protein Kinase Kinases