Overexpression Nanog Activates Pluripotent Genes in Porcine Fetal Fibroblasts and Nuclear Transfer Embryos

Zhang, Li; Luo, Yan; Bou, Gerelchimeg; Kong, Qingran; Ye, Huan; Zhu, Jiang; Wang, Yunlong; Li, Hui; Wang, Feng; Shi, Yuhua; Wei, Yanchang; Z, Liu

doi:10.1002/ar.21457

Cited by 22 publications

(19 citation statements)

References 39 publications

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“…It has been shown that overexpression of NANOG in donor cells had no effect on blastocyst rate in reconstructed embryos (Zhang et al, 2011). In our present study, similarly, the higher expression of NANOG in cloned AMSC-NT embryos could not improve the embryo development.…”

Section: Igf2 Igf2rcontrasting

confidence: 49%

The effect of amniotic membrane stem cells as donor nucleus on gene expression in reconstructed bovine oocytes

Nazari¹,

Shirazi²,

Shams‐Esfandabadi³

et al. 2016

Int. J. Dev. Biol.

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Nuclear reprogramming of a differentiated cell in somatic cell nuclear transfer (SCNT) is a major concern in cloning procedures. Indeed, the nucleus of the donor cell often fails to express the genes which are a prerequisite for normal early embryo development. This study was aimed to evaluate the developmental competence and the expression pattern of some reprogramming related genes in bovine cloned embryos reconstructed with amniotic membrane stem cells (AMSCs) in comparison with those reconstructed with mesenchymal stem cells (MSCs) and adult fibroblasts (AF) as well as with in vitro fertilized (IVF) oocytes. In vitro matured abattoir-derived oocytes were considered as recipients and a hand-made cloning technique was employed for oocyte enucleation and nuclear transfer (NT) procedures. The expression pattern of genes involved in self-renewal and pluripotency (POU5F1, SOX2, NANOG), imprinting (IGF2, IGF2R), DNA methylation (DNMT1, DNMT3A), histone deacetylation (HDAC2), and apoptosis (BAX, BCL2) were evaluated in NT and IVF derived embryos. Despite the insignificant difference in cleavage rate between reconstructed and IVF oocytes, the blastocyst rate in the IVF group was higher than that of other groups. Among reconstructed oocytes, a higher blastocysts rate was observed in MSC-NT and AMSCs-NT derived embryos that were significantly higher than AF-NT derived ones. There were more similarities in the expression pattern of pluripotency and epigenetic modification genes between MSC-NT and IVF derived blastocysts compared with other groups. In conclusion, considering developmental competence, AMSCs, as alternative donors in SCNT procedure, like MSCs, were prone to have more advantage compared with AF.

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Section: Igf2 Igf2rcontrasting

confidence: 49%

The effect of amniotic membrane stem cells as donor nucleus on gene expression in reconstructed bovine oocytes

Nazari¹,

Shirazi²,

Shams‐Esfandabadi³

et al. 2016

Int. J. Dev. Biol.

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“…Measurement of gene expression with quantitative real-time PCR has been applied in our previous research [19, 20]. Briefly, total RNA was extracted from 30 pooled embryos at each stage using an RNeasy Mini Kit (Qiagen, Dusseldorf, Germany) according to the manufacturer’s instructions.…”

Section: Methodsmentioning

confidence: 99%

Treating Cloned Embryos, But Not Donor Cells, with 5-aza-2’-deoxycytidine Enhances the Developmental Competence of Porcine Cloned Embryos

Huan

Zhu

Xie

et al. 2013

Journal of Reproduction and Development

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The efficiency of cloning by somatic cell nuclear transfer (SCNT) has remained low. In most cloned embryos, epigenetic reprogramming is incomplete, and usually the genome is hypermethylated. The DNA methylation inhibitor 5-aza-2’-deoxycytidine (5-aza-dC) could improve the developmental competence of cow, pig, cat and human SCNT embryos in previous studies. However, the parameters of 5-aza-dC treatment among species are different, and whether 5-aza-dC could enhance the developmental competence of porcine cloned embryos has still not been well studied. Therefore, in this study, we treated porcine fetal fibroblasts (PFF) that then were used as donor nuclei for nuclear transfer or fibroblast-derived reconstructed embryos with 5-aza-dC, and the concentration- and time-dependent effects of 5-aza-dC on porcine cloned embryos were investigated by assessing pseudo-pronucleus formation, developmental potential and pluripotent gene expression of these reconstructed embryos. Our results showed that 5-aza-dC significantly reduced the DNA methylation level in PFF (0 nM vs. 10 nM vs. 25 nM vs. 50 nM, 58.70% vs. 37.37% vs. 45.43% vs. 39.53%, P<0.05), but did not improve the blastocyst rate of cloned embryos derived from these cells. Treating cloned embryos with 25 nM 5-aza-dC for 24 h significantly enhanced the blastocyst rate compared with that of the untreated group. Furthermore, treating cloned embryos, but not donor cells, significantly promoted pseudo-pronucleus formation at 4 h post activation (51% for cloned embryos treated, 34% for donor cells treated and 36% for control, respectively, P<0.05) and enhanced the expression levels of pluripotent genes (Oct4, Nanog and Sox2) up to those of in vitro fertilized embryos during embryo development. In conclusion, treating cloned embryos, but not donor cells, with 5-aza-dC enhanced the developmental competence of porcine cloned embryos by promotion of pseudo-pronucleus formation and improvement of pluripotent gene expression.

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“…It has been shown that embryos cloned with highly expressed Oct4 cells enhanced blastocyst development, indicating that those cells underwent initial reprogramming events quite similar to normal fertilization (Pfeiffer et al, 2010). In embryos cloned with Nanog overexpression cells, Nanog overexpression did not have an effect on blastocyst rate or total cell number (Zhang et al, 2011).…”

Section: Introductionmentioning

confidence: 98%

Development and Gene Expression of Porcine Cloned Embryos Derived from Bone Marrow Stem Cells with Overexpressing Oct4 and Sox2

Lee

Lee²,

Jeon³

et al. 2014

Cellular Reprogramming

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The present study compared the potential of porcine bone marrow mesenchymal stem cells (pBMSCs) at different passages as nuclear transfer (NT) donors and the developmental efficiency of NT embryos from donor cells transfected with/without Oct4 and Sox2. Early-passage pBMSCs showed higher proliferation and expression of Oct4 and Sox2 and differentiation potential into mesenchymal lineages than middle- and late-passage pBMSCs. Cleavage rate did not differ among pBMSCs at different passages, but NT embryos with early-passage pBMSCs and middle-passage pBMSCs transfected with Oct4 (Oct4-pBMSCs) had significantly (p<0.05) higher blastocyst development than those with middle-passage pBMSCs. The incidence of apoptotic bodies in NT blastocysts from late-passage pBMSCs and Sox2-transfected middle-passage pBMSCs (Sox2-pBMSCs) was significantly (p<0.05) higher than others. The transcriptional levels of Oct4, Sox2, Nanog, Cdx2, Dnmt3a, and Igf2r genes were significantly (p<0.05) higher in Oct4- and Sox2-pBMSCs NT embryos. Middle-passage pBMSCs NT embryos revealed lower transcriptional levels of Bcl2 than others, except Sox2-pBMSCs NT embryos. The transcriptional level of Bax increased gradually in NT embryos derived from pBMSCs following extended passages and was significantly (p<0.05) higher in Sox2-pBMSCs NT embryos. Our results demonstrated that early-passage pBMSCs are more potent in expressing transcription factors and displayed higher differentiation ability, and middle-passage pBMSCs transfected with Oct4 improved the developmental efficiency of NT embryos, suggesting that high Oct4 expression cells are more efficient as NT donors.

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Overexpression Nanog Activates Pluripotent Genes in Porcine Fetal Fibroblasts and Nuclear Transfer Embryos

Cited by 22 publications

References 39 publications

The effect of amniotic membrane stem cells as donor nucleus on gene expression in reconstructed bovine oocytes

The effect of amniotic membrane stem cells as donor nucleus on gene expression in reconstructed bovine oocytes

Treating Cloned Embryos, But Not Donor Cells, with 5-aza-2’-deoxycytidine Enhances the Developmental Competence of Porcine Cloned Embryos

Development and Gene Expression of Porcine Cloned Embryos Derived from Bone Marrow Stem Cells with Overexpressing Oct4 and Sox2

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