• Journal of Embryo Transfer
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• v.21 no.1
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• pp.53-58
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• 2006

This study was conducted to comparing on vitrification of mouse oocytes and embryos using CPS, conventional straws and CPS by evaluating in morphological survival for oocytes, and embryonic cleavages and blastocyst formation for embryos. The morphological survival in vitro after thawing of vitrified oocytes using CPS (75%) and conventional straws (72%) were significantly higher (p<0.05) than that using OPS (68%). The blastocyst formation rates of vitrified embryos using CPS (48.6%) and unfrozen control embryos (56.0%) were significantly higher (p<0.05) than those of conventional straws (43.4%) and OPS (37.7%). The rates of morula formation were also higher to control, CPS, conventional straws and OPS in orderly. These results show that CPS has the advantages of achieving a high survival and safety preservation.

• Proceedings of the KSAR Conference
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• 2002.06a
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• pp.11-11
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• 2002

Interspecies nuclear transfer has been interested to determine ability of oocyte cytoplasm to support reprogramming of somatic cell nuclei of different species. In this study, we investigated developmental ability and mRNA expression patterns of developmentally important genes in bovine reconstructed embryos using a mouse fibroblast cell nucleus. While 20% nuclear transferred embryos with bovine fibroblast developed to morulae/blastocysts, a few(2-5%) nuclear transferred bovine embryos with mouse fibroblast developed to morula. (omitted)

• Korean Journal of Animal Reproduction
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• v.7 no.1
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• pp.24-29
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• 1983

These experiments were carried out to obtain basic information necessary of the success of in vitro culture of blastomeres separated from mouse embryo. Total 446 single blastomeres separated from 2-, 4- and 8-cell mouse embryos by protease treatment (0.5% in Whittingham's medium), were cultured under the gas phase of 5% CO2 in air at 37$^{\circ}C$. whittingham's medium was used for culture of blastomeres. The results obtained in these experiments were summerized as follows: 1. Of total 446 blastomeres cultured, 127(87.0%), 134(73.2%) and 77(65.8%) blastomeres separated repectively from 2-, 4- and 8-cell embryos were developed to morula or blastular stages. 2. The numbers of blastomeres, being separated from 2-. 4- and 8-cell embryos and developing to blastocysts containing inner cell mass, were 97(76.4%), 86(64.2%) and 33(42.9%) respectively. 3. After in vitro culture of the blastomeres, the incidence of trophoblastic vesicles increased with the development of the cell stage of embryo. In case of blastomeres separated from 8-cell embryos, 50.6% of blastomeres that developed to blastular stage was trophoblastic vesicles.

• Korean Journal of Animal Reproduction
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• v.16 no.4
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• pp.347-352
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• 1993

This study was designed to develop the in vitro culture systemof mammalian preimplantation embryos. We proposed mouse fetal fibroblast cells (MFFC) from 14∼15 day mouse fetus. Zygotes from superovulated female ICR mice were cultured 96 hrs in simple defined media (T6) or on the monolayer of MFFC. In addition, to evaluate the action of the co-culture of MFFC, various diluted superoxide dismutase antibody (SOD-Ab) was supplemented into the monolayer of MFFC and zygotes were cultrued in presence or absence of SOD-Ab. The developmental rates of zygotes were significantly increased in co-culture with MFFC compared to the control. The rates of zygotes to the 4-cell stage in media treated with EDTA were higher than those cultured in MFFC but the proportions of morula and blastocyst were not differ between EDTA and MFFC. Interestingly blastocysts in co-culture with MFFC possessed as many as blastomere as those developing in vivo, but blastocysts cultured with EDTA had significantly fewer blastomeres. In addition, the treatment of SOD-Ab suppressed the beneficial effect of MFFC. Therefore, our findings suggest that co-cultrue system using MFFC may have an advantage in the development of mouse zygotes as well as embryonic differentiation.

• Journal of Yeungnam Medical Science
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• v.12 no.1
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• pp.124-134
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• 1995

It is the most important to select optimal culture conditions to promote safe embryo growth in the technique of human in vitro fertilization and embryo transfer. It has been shown that the addition of biologic fluids, such as blood serum, of various origins, improved fertilization and early cleavage rates in numerous species. The purpose of this study is to attempt to measure developmental potential of mouse eggs fertilized and cleaved in Ham's F10 culture medium containing a chelating agent, EDTA and fetal cord serum. In this study, we selected 40 female mice and 20 male mice, and investigated optimal serum concentrations for mouse embryo growth. Two cell stage mouse embryos were cultured in Ham's F-10 medium, Ham's F-10 medium with various concentrations of EDTA, or Ham's F-10 medium with EDTA and 10% human cord serum. Developmental ratios to morula in Ham's F-10 medium containing various concentrations of EDTA and/or 10% fetal cord serum were significantly higher than in unsupplemented Ham's F-10 medium (p<0.05). Developmental ratios to blastocyst in Ham's F-10 containing 10% fetal cord serum and $50{\mu}M$ or $100{\mu}M$ EDTA were significanltly higher than in unsupplemented Ham's F-10 medium (p<0.05). Developmental ratios to morula in Ham's F-10 containing 10% fetal cord serum and $100{\mu}M$ EDTA were significanltly higher than in Ham's F-10 with 10% fetal cord serum used commonly in many human IVF centers(p<0.05). Developmental ratio to blastocyst in Ham's F-10 containing 10% fetal cord serum and $100{\mu}M$ EDTA was significanlty higher than in Ham's F-10 with $200{\mu}M$ EDTA(P<0.05). In summary, embryo development to morula and blastocyst was significanlty higher in the presence of human cord serum or EDTA than in the unsupplemented medium. The most significanly development to morula and blastocyst was obtained at Ham's F-10 medium with $100{\mu}M$ concentration of EDTA and 10% fetal cord serum. These results suggest that Ham's F-10 medium containing 10% fetal cord serum and optimal concentrations of EDTA significantly promoted early cleavage of mouse zygotes, and these will be useful as basic data for the selection of culture medium in human in vitro fertilization.

• Reproductive and Developmental Biology
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• v.31 no.1
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• pp.21-28
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• 2007

During early embryo development, Oct-4 is an important transcription factor for the early differentiation the present study was first examined methylation status in distal enhancer and promoter region of Oct-4 during mouse pre-implantation embryo development. In oocyte and sperm, high methylation was observed in both distal and proximal of promoter in Oct-4. Following fertilization relatively high methylation level remained until 8-cell stage embryos, but decreased at the morula and blastocyst stage. Specific gene knock down of Oct-4 by siRNA injection into zygote induced higher methylation rates of both distal and proximal region of promoter of Oct-4. These results suggest a functional link between the DNA methylation status of distal and promoter resign in the Oct-4 gene and the gene sequence-specific transcriptional silencing by exogenous siRNA injection during mouse preimplantation embryos.

• Korean Journal of Animal Reproduction
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• v.13 no.3
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• pp.141-148
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• 1989

The effects of seeding method and optimum time for freezing embryos according to the developmental stages on embryo survival rates after rapid freezing were determined using the FDA-test. The summarized results are as follows : 1. In the rapid freezing of embryos, the sucrose added medium together with Co-seeding or non-seeding showed the FDA scores of 4.67 and 4.20, respectively, but, raffinose addition obtained FDA scores of 4.27 and 3.97. 2. The developmental stage of embryos at freezing was most critical on the survival of embryos after thawing. Higher FDA scores were obtained in the order of blastocyst stage(4.94), morula stage(3.82) and ealy stage(2.65) in sucrose added medium. The same trend was observed in the raffinose added medium with an order or 4.91, 4.47 and 2.32. 3. Microscopic study of embryo before freezing and post-thawing indicated that the embryo showed shrinkage within 5 minutes after the embryo was transfer to the freezing medium. When thawed embryo was tranfered to the dilution medium, swelling of the embryo was observed and there after it reshrank indicating the removal of cryoprotectant from the embryo. The size of the embryo recovered to the original state when it was moved into a PBS-solution.

• Development and Reproduction
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• v.5 no.1
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• pp.17-21
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• 2001

This study was conducted to investigate the expression pattern of Trypsin-like enzyme and the effect of a trypsin inhibitor(benzimidine) on hatching process during in-vitro culture of mouse preimplantation embryos. The Trypsin-like enzyme was identified by rhodamine-conjugated Trypsin substrate probe. The expression of trypsin-like enzyme was firstly detected at the late morula stage, and the enzyme was uniformly localized in the trophectoderm of late blastocysts. Especially, intense fluorescence was observed in the blebbing area of hatching blastocysts. Bisbenzamidine, contained in culture media, did not alter embryonic development from 4-cell stage to the expanded blastocyst but decrease the hatching rate in ImM concentration (15.8% vs 89.7%, p<0.02). In the treatment of bisbenzimidine (5mM) for 12 hours according to the embryonic stage of mouse, the hatching rate of control (83.0%) and treatment in late blastocysts (8.7%) were significantly (p<0.01) different. From these results, we suggested that the hatching enzyme having trypsin-like activity was localized from the late morula stage, and the hatching process by this enzyme was activated in the late blastocyst stage of mouse embryos.

• Journal of Embryo Transfer
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• v.8 no.1
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• pp.31-36
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• 1993

The post-thaw survival of mouse embryos of the various developmental stages was determined after cryopreservation by vitrification in a solution containing ethylene glycol, Ficoll and sucrose (EFS). All the embryos were equilibrated for 2 minutes just prior to freezing. The number of blastomeres during in vitro development was counted by nuclei higher rates of post-thaw survival were obtained from the embryos of 2-cell(92.2%), 8-cell(77.2%) or morula stage(90.0%) than those of blastocyst stage(62.7%). The number of blastomeres per embryo following in vitro culture for 24 hours was significantly(P<0.05) smaller as 66.0f22.3 in vitrified and thawed morulae than fresh morulae(91.7$\pm$12.2).

• Animal cells and systems
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• v.10 no.1
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• pp.41-47
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• 2006

Development of the mammalian pre-implantation embryos has unique features, such as a slow and unsynchronized cell division, compaction, and eventual formation of blastocysts with inner cell mass and trophectoderm. In order to have a clue on molecular mechanisms that reside in mouse early development, we suppressed expression of early embryo-specific genes with RNAi and observed their development in vitro. We observed developmental defects in embryos microinjected with dsRNAs for Oct4 or Nanog among the tested genes. Careful examinations revealed that development of the most of the Oct4- or Nanog-suppressed embryos were arrested at the morula stage. These results suggest that the Oct4 and Nanog activities are also required for embryogenesis earlier than the blastocyst stage.