scholarly journals Nuclear Transfer of Embryonic Cell Nuclei to Non-enucleated Eggs in Zebrafish, Danio rerio

2011 ◽  
Vol 7 (4) ◽  
pp. 460-468 ◽  
Author(s):  
Manabu Hattori ◽  
Hisashi Hashimoto ◽  
Ekaterina Bubenshchikova ◽  
Yuko Wakamatsu
Keyword(s):  
Danio Rerio ◽  
Nuclear Transfer ◽  
Embryonic Cell ◽  
Cell Nuclei

Differential ability of male and female rabbit fetal germ cell nuclei to be reprogrammed by nuclear transfer

1996 ◽  
Vol 60 (5) ◽  
pp. 339-345 ◽  
Author(s):  
André Moens ◽  
Sylvie Chastant ◽  
Patrick Chesné ◽  
Jacques-Edmond Fléchon ◽  
Keith J. Betteridge ◽  
...  
Keyword(s):  
Germ Cell ◽  
Nuclear Transfer ◽  
Cell Nuclei ◽  
Male And Female ◽  
Female Rabbit ◽  
Differential Ability ◽  
Fetal Germ Cell

62 REPROGRAMMING EVENTS AND DEVELOPMENTAL COMPETENCE OF RHESUS MONKEY EMBRYOS PRODUCED BY SOMATIC CELL NUCLEAR TRANSFER

2006 ◽  
Vol 18 (2) ◽  
pp. 139 ◽  
Author(s):  
S. Mitalipov ◽  
Q. Zhou ◽  
J. Byrne ◽  
W.-Z. Ji ◽  
D. Wolf
Keyword(s):  
Somatic Cell ◽  
Somatic Cell Nuclear Transfer ◽  
Nuclear Transfer ◽  
Formation Rate ◽  
Embryonic Stem ◽  
Developmental Competence ◽  
Lamin A ◽  
Cell Nuclei ◽  
Blastocyst Development ◽  
Morula Stage

Successful reprogramming of somatic cell nuclei after nuclear transfer requires active remodeling by factors present in the nonactivated cytoplast. High levels of maturation promoting factor (MPF) activity are associated with this remodeling process which includes nuclear envelope breakdown (NEBD), premature chromosome condensation (PCC), and spindle formation. In this study, we examined the extent of nuclear remodeling in monkey somatic cell nuclear transfer (SCNT) embryos by monitoring the dynamics of lamin A/C appearance, as detected immunocytochemically, following fusion of donor cells with recipient cytoplasts. In the control, intracytoplasmic sperm injection (ICSI) fertilized embryos, lamin A/C was readily detected at the pronuclear stage but disappeared in early cleaving embryos only to reappear by the morula stage in association with the activation of the embryonic genome. We initially documented lack or incomplete NEBD and PCC in SCNT embryos in the form of retention of lamin A/C signal emanating from the donor nucleus. This observation was consistent with premature cytoplast activation due to the manipulation procedures. SCNT embryos produced by this approach typically arrested at the morula stage. Significant modifications in nuclear transfer protocols were then employed. Optimization of procedures resulted in robust NEBD and PCC, as indicated by loss of lamin A/C signal from the donor cell. Also, significant improvement of SCNT embryo development in vitro was observed, with a markedly improved blastocyst formation rate (21%). Several different fetal and adult somatic cell types screened as nuclear donors supported blastocyst development. SCNT blastocysts displayed a pattern of Oct-4 expression similar to that of sperm fertilized counterparts, indicative of efficient nuclear reprogramming. However, no pregnancies were established following a preliminary trial of 8 embryo transfers with 48 cloned embryos. Nevertheless, our results represent a breakthrough in efforts to produce cloned monkeys and should provide the resources required for the derivation of embryonic stem cells from SCNT blastocysts.

47 DIFFERENTIAL DEVELOPMENTAL ABILITY OF EMBRYOS CLONED FROM TISSUE-SPECIFIC STEM CELLS

2007 ◽  
Vol 19 (1) ◽  
pp. 142
Author(s):  
K. Inoue ◽  
N. Ogonuki ◽  
H. Miki ◽  
S. Noda ◽  
S. Inoue ◽  
...  
Keyword(s):  
Stem Cells ◽  
Nuclear Transfer ◽  
Embryonic Stem ◽  
Donor Cell ◽  
Fibroblast Cell ◽  
High Rate ◽  
Full Potential ◽  
Cell Nuclei ◽  
Cell Stage

Although cloning animals by somatic cell nuclear transfer is generally an inefficient process, use of appropriate donor cell types may improve the cloning outcome significantly. Among the donor cells tested so far, mouse embryonic stem cells have given the best efficiency in terms of the development of reconstructed embryos into offspring. In this study, we examined whether 2 in vitro-produced pluripotent stem cells—neural stem cells (NSCs) and mesenchymal stem cells (MSCs)—could be better nuclear donors than other differentiated cells. Embryos were reconstructed by transfer of nuclei from NSCs or MSCs with full potential for differentiation in vitro. Most (76%) of the 2-cell NCS embryos developed to the 4-cell stage; 43% implanted and 1.6% developed to term after transfer to pseudopregnant recipients. These rates were very similar to those of embryos cloned from fibroblast cell nuclei. Interestingly, in the patterns of zygotic gene expression, NSC embryos were more similar to in vitro-fertilized embryos than fibroblast cloned embryos. By contrast, embryos reconstructed using MSC nuclei showed lower developmental ability and no implantation was obtained after embryo transfer. Chromosomal analysis of the donor MSCs revealed very high frequencies of monosomy and trisomy, which might have caused the very poor post-implantation development of embryos following nuclear transfer. Thus, in vitro-produced pluripotent cells can serve as donors of nuclei for cloning mice, but may be prone to chromosomal aberrations leading to a high rate of cloned embryo death.

Micromanipulation medium osmolarity compromises zebrafish (Danio rerio) embryo and cell survival in chimaerism experiments

Zygote ◽  
2009 ◽  
Vol 18 (2) ◽  
pp. 155-158 ◽  
Author(s):  
J. Cardona-Costa ◽  
M. Francisco-Simão ◽  
M. Pérez-Camps ◽  
F. García-Ximénez
Keyword(s):  
Danio Rerio ◽  
External Medium ◽  
Mechanical Damage ◽  
Donor Cell ◽  
Embryonic Cell ◽  
Zebrafish Embryos ◽  
Embryo Survival ◽  
Cell Injection ◽  
Donor Cells ◽  
Medium Osmolarity

SummaryIn zebrafish chimaerism experiments, the cell injection can involve intra-embryonic cell lyses by osmolar effects. Moreover, the donor cells can be injured during manipulation due to osmolar changes into the transplant pipette. Therefore, the present study aimed to assess the effects of manipulation medium osmolarity on embryonic survival and donor cell viability.In Experiment I, 0.1 μl to 0.15 μl approximately of an isosmolar solution (300 mOsm) was injected into recipient embryos, which were kept at 300 (E1) or 30 mOsm (E2). Survival at day 1 was significantly higher in the E2 group than in E1 (E1: 68% vs E2: 81%, p < 0.05), but after 5 days embryo survival in the E1 group was slightly higher. In Experiment II, donor cells from zebrafish embryos were exposed (or not) to a possible osmolarity change (inner pipette medium: 300 mOsm vs external medium: 30 or 300 mOsm) using two different micropipette outer diameters, 40–50 and 60–70 μm. Cell mechanical damage was detected in the 40–50 μm pipette (p < 0.05), but not by the handling medium osmolarity. Results recommend the use of a 300 mOsm manipulation medium and bore-sized pipettes adjusted as closely as possible to the donor cell size.

scholarly journals Bovine Oocyte Cytoplasm Supports Development of Embryos Produced by Nuclear Transfer of Somatic Cell Nuclei from Various Mammalian Species1

1999 ◽  
Vol 60 (6) ◽  
pp. 1496-1502 ◽  
Author(s):  
Tanja Dominko ◽  
Maissam Mitalipova ◽  
Brad Haley ◽  
Zeki Beyhan ◽  
Erdogan Memili ◽  
...  
Keyword(s):  
Somatic Cell ◽  
Nuclear Transfer ◽  
Bovine Oocyte ◽  
Cell Nuclei ◽  
Oocyte Cytoplasm

scholarly journals Production of Nuclear Transfer Horse Embryos by Piezo-Driven Injection of Somatic Cell Nuclei and Activation with Stallion Sperm Cytosolic Extract1

2002 ◽  
Vol 67 (2) ◽  
pp. 561-567 ◽  
Author(s):  
Y.H. Choi ◽  
C.C. Love ◽  
Y.G. Chung ◽  
D.D. Varner ◽  
M.E. Westhusin ◽  
...  
Keyword(s):  
Somatic Cell ◽  
Nuclear Transfer ◽  
Cell Nuclei

47 SUCCESSFULLY PRODUCING CLONED MICE FROM SOMATIC CELLS OF AGED MICE VIA ESTABLISHED ntES CELL LINES

2008 ◽  
Vol 20 (1) ◽  
pp. 104
Author(s):  
E. Mizutani ◽  
T. Ono ◽  
L. Chong ◽  
T. Wakayama
Keyword(s):  
Somatic Cell ◽  
Cell Lines ◽  
Nuclear Transfer ◽  
Somatic Cells ◽  
Chimeric Mice ◽  
Cell Nuclei ◽  
Female Mice ◽  
Aged Mice ◽  
Tail Tip ◽  
Tip Cells

Recent nuclear transfer techniques have enabled us to produce cloned animals from somatic cell nuclei in a variety of animal species and are to date the only way to obtain offspring from infertile animals. Despite very aged mice often showing an infertile phenotype, the decreasing rate of cloning success with increased age makes it almost impossible to produce cloned mice or offspring from these animals. Other studies, however, have demonstrated that ES cell lines have been established from cloned blastocysts through somatic cell nuclear transfer (ntES cells), irrespective of sex, strains, or organs. These cells are subsequently capable of differentiating into all three germ layers in vitro, or even into spermatozoa and oocytes in chimeric mice. Thus, ntES cells have received considerable attention recently in regenerative medicine. Importantly, the success rate of establishing ntES cell lines from cloned blastocysts is ten times higher than that of producing cloned mice, which may allow us to establish ntES cell lines even from such 'unclonable' aged mice. ntES cells also have the potential to be a good donor source for nuclear transfer as they have the same DNA as their donor somatic cells and can indefinitely proliferate in their undifferentiated states. In this study, we attempted to establish ntES cell lines from aged mice and analyze their normality. We then tried to produce cloned mice via nuclear transfer using established ntES cell lines. We obtained donor cells from tail-tip fibroblast cells of BDF1 and BCF1 male and female mice that were over two years old. Following nuclear transfer, we transferred a proportion of the cleaved cloned embryos to pseudopregnant ICR female mice. The remaining embryos were cultured for 72 h, and cloned embryos that developed into morulae or blastocysts were plated on feeder cells. We then examined all established ntES cell lines for normality by Oct4 and Nanog expression using immunofluorescence staining and pluripotency by chimeric mice formation, for which ntES cells were injected into fertilized ICR embryos. Finally, we attempted to produce cloned mice from the nuclei of these ntES cell lines. In each experiment, 25, 37, 73, 63, and 75 cloned embryos from aged mice tail-tip cells were used in attempts to produce cloned mice, and 20, 20, 27, 35, and 40 cloned embryos were used to derive ntES cell lines, respectively. No cloned mice were obtained by direct nuclear transfer of the aged mice tail-tip cells; however, we were successful in establishing ntES cell lines from all experiments, with an establishment rate between 10 and 25%. All established ntES cell lines expressed Oct4 and Nanog and contributed to somatic cells in chimeric mice. Some chimeric mice produced offspring derived from ntES cells after mating. We were also able to produce cloned mice even from a 2-year-old and a 9-month-old BCF1 male mouse by nuclear transfer using ntES cells as donor nuclei. These results clearly show that normal ntES cell lines can be established from infertile, aged mice and this technique can now be used to produce offspring, irrespective of donor conditions.

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