Long-term effect on in vitro cloning efficiency after treatment of somatic cells with Xenopus egg extract in the pig

Ying Liu; Olga Østrup; Rong Li; Juan Li; Gabor Vajta; Peter Michael Kragh; Mette Schmidt; Stig Purup; Poul Hyttel; Dan Arne Klærke; Henrik Callesen

doi:10.1071/rd13147

Long-term effect on in vitro cloning efficiency after treatment of somatic cells with Xenopus egg extract in the pig

Ying Liu, Olga Østrup, Rong Li, Juan Li, Gabor Vajta, Peter Michael Kragh, Mette Schmidt, Stig Purup, Poul Hyttel, Dan Arne Klærke, Henrik Callesen

7 Citations (Scopus)

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.

Original language	English
Journal	Reproduction, Fertility and Development
Volume	26
Issue number	7
Pages (from-to)	1017-1031
Number of pages	15
ISSN	1031-3613
DOIs	https://doi.org/10.1071/rd13147
Publication status	Published - 2014

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@article{766463bea5c14fb288ee7cf753ef88a0,

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.",

author = "Ying Liu and Olga {\O}strup and Rong Li and Juan Li and Gabor Vajta and Kragh, {Peter Michael} and Mette Schmidt and Stig Purup and Poul Hyttel and Kl{\ae}rke, {Dan Arne} and Henrik Callesen",

year = "2014",

doi = "10.1071/rd13147",

language = "English",

volume = "26",

pages = "1017--1031",

journal = "Reproduction, Fertility and Development",

issn = "1031-3613",

publisher = "C S I R O Publishing",

number = "7",

}

TY - JOUR

T1 - Long-term effect on in vitro cloning efficiency after treatment of somatic cells with Xenopus egg extract in the pig

AU - Liu, Ying

AU - Østrup, Olga

AU - Li, Rong

AU - Li, Juan

AU - Vajta, Gabor

AU - Kragh, Peter Michael

AU - Schmidt, Mette

AU - Purup, Stig

AU - Hyttel, Poul

AU - Klærke, Dan Arne

AU - Callesen, Henrik

PY - 2014

Y1 - 2014

N2 - 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.

AB - 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.

U2 - 10.1071/rd13147

DO - 10.1071/rd13147

M3 - Journal article

C2 - 25145414

SN - 1031-3613

VL - 26

SP - 1017

EP - 1031

JO - Reproduction, Fertility and Development

JF - Reproduction, Fertility and Development

IS - 7

ER -

Long-term effect on in vitro cloning efficiency after treatment of somatic cells with Xenopus egg extract in the pig

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