Long-term effect on in vitro cloning efficiency after treatment of somatic cells with Xenopus egg extract in the pig
Ying Liu A E , Olga Østrup B , Rong Li A , Juan Li A C , Gábor Vajta A , Peter M. Kragh A , Mette Schmidt D , Stig Purup A , Poul Hyttel B , Dan Klærke B and Henrik Callesen A
+ Author Affiliations
- Author Affiliations
A Department of Animal Science, Aarhus University, DK-8830 Tjele, Denmark.
B Department of Veterinary Clinical and Animal Sciences, University of Copenhagen, DK-1870 Frederiksberg C, Denmark.
C College of Animal Science, Nanjing Agricultural University, Nanjing Weigang No.1, 210095, China.
D Department of Veterinary Reproduction and Obstetrics, University of Copenhagen, DK-1870 Frederiksberg C, Denmark.
E Corresponding author. Email: ying.liu@agrsci.dk
Reproduction, Fertility and Development 26(7) 1017-1031 https://doi.org/10.1071/RD13147
Submitted: 12 May 2013 Accepted: 2 July 2013 Published: 8 August 2013
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.
Additional keywords: colony, embryo development, reprogramming, somatic cell nuclear transfer.
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