<i>LacZ</i> transgene expression as a cell marker to analyse rescue from the 2-cell block in mouse aggregation chimeras

Neganova, Irina; Augustin, Martin; Sekirina, Galina G.; Jockusch, Harald

doi:10.1017/s096719949800015x

Cited by 9 publications

(9 citation statements)

References 10 publications

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“…In contrast to the mouse data, a recent report in bovine showed no improvement in the development rate of NT aggregates or OCT4 levels when compared to non-aggregates, although the cell number in aggregates was increased (Misica-Turner et al, 2006). This apparent discrepancy and early studies (Sekirina and Neganova, 1995;Neganova et al, 1998) suggest that beneficial complementation between aggregated cells may be regulated by the timing of the aggregation. Mouse NT aggregation was performed after the onset of major embryonic genome activation, while up to now all the bovine NT aggregations have been conducted before genome activation.…”

Section: Discussionmentioning

confidence: 69%

Global gene expression analysis of bovine blastocysts produced by multiple methods

Zhou¹,

Xiang²,

Walker³

et al. 2007

Molecular Reproduction Devel

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Reproductive efficiency using somatic cell nuclear transfer (SCNT) technology remains suboptimal. Of the various efforts to improve the efficiency, chromatin transfer (CT) and clone-clone aggregation (NTagg) have been reported to produce live cloned animals. To better understand the molecular mechanisms of somatic cell reprogramming during SCNT and assess the various SCNT methods on the molecular level, we performed gene expression analysis on bovine blastocysts produced via standard nuclear transfer (NT), CT, NTagg, in vitro fertilization (IVF), and artificial insemination (AI), as well as on somatic donor cells, using bovine genome arrays. The expression profiles of SCNT (NT, CT, NTagg) embryos were compared with IVF and AI embryos as well as donor cells. NT and CT embryos have indistinguishable gene expression patterns. In comparison to IVF or AI embryos, the number of differentially expressed genes in NTagg embryos is significantly higher than in NT and CT embryos. Genes that were differentially expressed between all the SCNT embryos and IVF or AI embryos are identified. Compared to AI embryos, more than half of the genes found deregulated between SCNT and AI embryos appear to be the result of in vitro culture alone. The results indicate that although SCNT methods have altered differentiated somatic nuclei gene expression to more closely resemble that of embryonic nuclei, combination of insufficient reprogramming and in vitro culture condition compromise the developmental potential of SCNT embryos. This is the first set of comprehensive data for analyzing the molecular impact of various nuclear transfer methods on bovine pre-implantation embryos.

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Section: Discussionmentioning

confidence: 69%

Global gene expression analysis of bovine blastocysts produced by multiple methods

Zhou¹,

Xiang²,

Walker³

et al. 2007

Molecular Reproduction Devel

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“…Methylation Reprogramming Embryos from most inbred mouse strains or from hybrids between inbred strains, i.e., B6C3F1, can efficiently develop in vitro to the blastocyst stage. However, when embryos from NMRI or other so-called blocking strains are placed in culture before they have reached the mid two-cell stage, they will usually arrest development in the G2-phase of the second cell cycle (Monk, 1987;Neganova et al, 1998). This effect was particularly evident, when NMRI embryos were cultured in M16-medium without complements.…”

Section: Strain-specific Differences In the Efficiency Ofmentioning

confidence: 99%

Aberrant methylation patterns at the two‐cell stage as an indicator of early developmental failure

Wei

Haaf²

2002

Molecular Reproduction Devel

234

141

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The fertilized mouse egg actively demethylates the paternal genome within a few hours after fertilization, whereas the maternal genome is only passively demethylated by a replication-dependent mechanism after the two-cell stage. This evolutionarily conserved assymetry in the early diploid mammalian embryo may have a role in methylation reprogramming of the two very different sets of sperm and egg chromatin for somatic development and formation of totipotent cells. Immunofluorescence staining with an antibody against 5-methylcytosine (MeC) showed that the incidence of abnormal methylation patterns differs between mouse two-cell embryos from superovulated females, nonsuperovulated matings, and in vitro fertilization (IVF). It also depends on embryo culture conditions and genetic background. In general, there was a good correlation with the number of embryos (from the same experiment) which did not develop in vitro up to the blastocyst stage. Thus, aberrant genome-wide DNA methylation in early embryos may be an important mechanism contributing to the high incidence of developmental failure in mammals. Similar to the situation in abnormally methylated embryos from nuclear transfer, it may cause a high incidence of pregnancy loss and abnormal phenotypes.

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“…It has been suggested that the 2-cell block and other forms of early developmental failure or early pregnancy loss in mouse correlate with aberrant methylation patterns or methylation reprogramming defects (Li et al ., 1992; Neganova et al ., 1998; Patkin et al ., 1998; Shi & Haaf, 2002). DNA methylation belongs in the group of epigenetic modifications, which mark genomic regions and act as heritable and stable instructions for the specification of chromatin organization and structure, which in turn dictate transcriptional states.…”

Section: Introductionmentioning

confidence: 99%

Apoptotic processes and DNA cytosine methylation in mouse embryos arrested at the 2-cell stage

Fabian

Bukovská

Juhás

et al. 2009

Zygote

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The present study evaluates the role of apoptotic cell death and DNA methylation reprogramming in early developmental failures occurring in embryos at the 2-cell stage. Mouse 2-cell embryos were cultured in vitro and treated with chemicals that cause developmental arrest and apoptosis (alpha-amanitin, actinomycin D, TNF-alpha). After 24 h, 48 h and 72 h culture, embryos were analysed using cell-death assays (annexin V staining, TUNEL labelling and immunodetection of active caspase-3) and genome methylation assay (immunodetection of 5-methylcytosine). The ability of embryos at the 2-cell stage to undergo apoptotic processes was very low. In arrested embryos, the presence of all evaluated features of apoptosis was recorded only after 72 h culture and their incidence was sporadical. Interestingly, the most frequently observed apoptotic sign was nuclear condensation and the timing of its appearance preceded even the phosphatidylserine flip. Both normally developing and arrested embryos displayed reduction in DNA cytosine methylation. In arrested embryos, this process was independent of cellular cleavage, was more pronounced and finished in almost complete demethylation of the embryonic genome. The timing of the demethylation overlapped with the onset of major apoptotic events. Although observed apoptotic cells showed either demethylated or methylated DNA cytosine in their nuclei, at blastocyst stage the demethylated status appeared more frequently in them.

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LacZ transgene expression as a cell marker to analyse rescue from the 2-cell block in mouse aggregation chimeras

Cited by 9 publications

References 10 publications

Global gene expression analysis of bovine blastocysts produced by multiple methods

Global gene expression analysis of bovine blastocysts produced by multiple methods

Aberrant methylation patterns at the two‐cell stage as an indicator of early developmental failure

Apoptotic processes and DNA cytosine methylation in mouse embryos arrested at the 2-cell stage

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