title: Long-term effect on in vitro cloning efficiency after treatment of somatic cells with Xenopus egg extract in the pig.
abstract: In somatic cell nuclear transfer (SCNT), donor cell reprogramming is considered as a biologically important and vulnerable event. Various donor cell pre-treatments with Xenopus egg extracts can promote reprogramming. Here we investigated if the reprogramming effect of one treatment with Xenopus egg extract on donor cells was maintained for several cell passages. The extract treatment resulted in increased cell-colony formation from early passages in treated porcine fibroblasts (ExTES), and increased development of cloned embryos. Partial dedifferentiation was observed in ExTES cells, shown as a tendency towards upregulation of NANOG, c-MYC and KLF-4 and downregulation of DESMIM compared with ExTES at Passage 2. Compared with our routine SCNT, continuously increased development of cloned embryos was observed in the ExTES group, and ExTES cloned blastocysts displayed hypermethylated DNA patterns and hypermethylation of H3K4me3 and H3K27me3 in ICM compared with TE. All seven recipients became pregnant after transferral of ExTES cloned embryos and gave birth to 7-22 piglets per litter (average 12). In conclusion, our results demonstrate that one treatment of porcine fibroblasts with Xenopus egg extract can result in long-term increased ability of the cells to promote their in vitro function in subsequent SCNT. Finally these cells can also result in successful development of cloned embryos to term.
authors.affiliation: Department of Animal Science, Aarhus University, DK-8830 Tjele, Denmark.
authors.forename: Ying
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Department of Veterinary Clinical and Animal Sciences, University of Copenhagen, DK-1870 Frederiksberg C, Denmark.
authors.forename: Olga
authors.initials: O
authors.lastname: Ostrup
authors.affiliation: Department of Animal Science, Aarhus University, DK-8830 Tjele, Denmark.
authors.forename: Rong
authors.initials: R
authors.lastname: Li
authors.affiliation: Department of Animal Science, Aarhus University, DK-8830 Tjele, Denmark.
authors.forename: Juan
authors.initials: J
authors.lastname: Li
authors.affiliation: Department of Animal Science, Aarhus University, DK-8830 Tjele, Denmark.
authors.forename: Gábor
authors.initials: G
authors.lastname: Vajta
authors.affiliation: Department of Animal Science, Aarhus University, DK-8830 Tjele, Denmark.
authors.forename: Peter M
authors.initials: PM
authors.lastname: Kragh
authors.affiliation: Department of Veterinary Reproduction and Obstetrics, University of Copenhagen, DK-1870 Frederiksberg C, Denmark.
authors.forename: Mette
authors.initials: M
authors.lastname: Schmidt
authors.affiliation: Department of Animal Science, Aarhus University, DK-8830 Tjele, Denmark.
authors.forename: Stig
authors.initials: S
authors.lastname: Purup
authors.affiliation: Department of Veterinary Clinical and Animal Sciences, University of Copenhagen, DK-1870 Frederiksberg C, Denmark.
authors.forename: Poul
authors.initials: P
authors.lastname: Hyttel
authors.affiliation: Department of Veterinary Clinical and Animal Sciences, University of Copenhagen, DK-1870 Frederiksberg C, Denmark.
authors.forename: Dan
authors.initials: D
authors.lastname: Klærke
authors.affiliation: Department of Animal Science, Aarhus University, DK-8830 Tjele, Denmark.
authors.forename: Henrik
authors.initials: H
authors.lastname: Callesen
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D065150:Cellular Reprogramming / Q000235:genetics; D019976:Cloning, Organism / Q000379:methods / Q000662:veterinary*; D019175:DNA Methylation; D004624:Embryo Transfer / Q000662:veterinary; D047108:Embryonic Development; D005260:Female; D005347:Fibroblasts / Q000648:ultrastructure; D015870:Gene Expression; D006657:Histones / Q000378:metabolism; D008297:Male; D008745:Methylation; D009687:Nuclear Proteins / Q000032:analysis; D053652:Nuclear Transfer Techniques / Q000662:veterinary*; D000090243:Nucleophosmin; D010063:Ovum / Q000737:chemistry*; D035463:Pol1 Transcription Initiation Complex Proteins / Q000032:analysis; D011247:Pregnancy; D034421:Sus scrofa; D014981:Xenopus
chemical_list: D006657:Histones; D009687:Nuclear Proteins; D035463:Pol1 Transcription Initiation Complex Proteins; C063844:transcription factor UBF; D000090243:Nucleophosmin