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Published on: April 26, 2018
Effect of donor cell type on nuclear remodelling in rabbit somatic cell nuclear transfer embryos
1College of Animal Science and Veterinary Medicine, Jilin University, Changchun, Jilin, China.
Abstract:
Cloned rabbits have been produced for many years by somatic cell nuclear transfer (SCNT). The efficiency of cloning by SCNT, however, has remained extremely low. Most cloned embryos degenerate in utero, and the few that develop to term show a high incidence of post-natal death and abnormalities. The cell type used for donor nuclei is an important factor in nuclear transfer (NT). As reported previously, NT embryos reconstructed with fresh cumulus cells (CC-embryos) have better developmental potential than those reconstructed with foetal fibroblasts (FF-embryos) in vivo and in vitro. The reason for this disparity in developmental capacity is still unknown. In this study, we compared active demethylation levels and morphological changes between the nuclei of CC-embryos and FF-embryos shortly after activation. Anti-5-methylcytosine immunofluorescence of in vivo-fertilized and cloned rabbit embryos revealed that there was no detectable active demethylation in rabbit zygotes or NT-embryos derived from either fibroblasts or CC. In the process of nuclear remodelling, however, the proportion of nuclei with abnormal appearance in FF-embryos was significantly higher than that in CC-embryos during the first cell cycle. Our study demonstrates that the nuclear remodelling abnormality of cloned rabbit embryos may be one important factor for the disparity in developmental success between CC-embryos and FF-embryos.
Insights
Somatic cell nuclear transfer (SCNT) in rabbits has low efficiency. This study found that abnormal nuclear remodeling in fibroblast-derived cloned rabbit embryos, not demethylation, contributes to developmental failure.
Area of Science:
- Reproductive biology
- Developmental biology
- Animal cloning
Background:
- Somatic cell nuclear transfer (SCNT) is used for rabbit cloning but has low efficiency.
- Most cloned embryos degenerate or result in abnormal offspring.
- The donor cell type impacts cloning success, with cumulus cells (CC) outperforming fetal fibroblasts (FF).
Purpose of the Study:
- To investigate the reasons for developmental disparities between CC- and FF-derived cloned rabbit embryos.
- To compare active demethylation and nuclear remodeling in early-stage cloned embryos.
Main Methods:
- Somatic cell nuclear transfer (SCNT) using cumulus cells (CC) and fetal fibroblasts (FF) in rabbits.
- Immunofluorescence staining for 5-methylcytosine to assess DNA demethylation.
- Microscopic analysis of nuclear morphology during the first cell cycle post-activation.
Main Results:
- No significant active DNA demethylation was detected in rabbit zygotes or SCNT embryos (CC- or FF-derived).
- A higher proportion of abnormal nuclear morphology was observed in FF-derived cloned embryos compared to CC-derived embryos during the first cell cycle.
- Nuclear remodeling abnormalities were identified as a key difference between the two groups.
Conclusions:
- Active DNA demethylation is not the primary factor limiting rabbit cloning efficiency with these donor cell types.
- Nuclear remodeling abnormalities in the early stages of development are a significant factor contributing to the lower developmental success of cloned rabbit embryos derived from fetal fibroblasts.
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Somatic Cell Nuclear Transfer
In SCNT, an egg cell is taken from an animal and its nucleus is removed, creating an enucleated egg. Then a somatic cell—any cell that is not a sex...