• Asian-Australasian Journal of Animal Sciences
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• v.19 no.2
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• pp.171-175
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• 2006

The aim of this in vitro study was to test the effect of microinjection (Mi) of foreign gene into the rabbit egg pronucleus and epidermal growth factor (EGF) addition on the blastocyst rate, the cell number and the diameter of embryos, and to determine possible relationships between embryo cell number and embryo diameter. Blastocyst rate was significantly decreased in gene- Mi (G-Mi/E0) group (63.1%) comparing to intact ones (83.5%, $p_1$<0.05). The addition of EGF at 20ng/ml (G-Mi/E20) or 200 ng/ml (GMi/ E200) to gene-Mi embryos did not affect blastocyst rate (65.6 and 55.2% resp.). As a control for Mi, the eggs were microinjected with the same volume of phosphate-buffered solution (PBS-Mi) instead of the gene construct solution. Cell numbers and embryo diameters were measured from embryo images obtained on confocal laser scanning microscope. Bonferroni-modified LSD test showed that the embryo cell number in PBS-Mi group was significantly lower ($p_1$<0.05) and in gene-Mi group was tended to decrease compared with intact embryos. Embryo diameter was not different among experimental groups. No effect of EGF given at any doses both on the cell number and embryo diameter was found. A positive correlation between cell number and embryo diameter was observed in all groups of embryos. Since embryo diameter was not changed under the influence of Mi or EGF addition in this study, this seems to be more conservative characteristics of the embryo morphology. These results suggest that the pronuclear microinjection compromises developmental potential of embryos, decreasing blastocyst rate and embryo cell number, whilst embryo diameter is not affected. No effects of EGF on studied parameters were confirmed. Declined quality of Mi-derived embryos is caused by the microinjection procedure itself, rather than by the gene construct used.

• Korean Journal of Animal Reproduction
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• v.21 no.2
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• pp.85-93
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• 1997

These experiments were carried out to examine the development capacity of sexed and then bisected embryo from 8-cell to morula stage. Antisera to histocompatibility-Y(H-Y) antigen were prepared in inbred SD female rat by repeated immunization of spleen cell or testis supernatant from males of same strain. Male and female embryos were separated by delaying development of embryos against H-Y antibody. After sexing, rabbit embryos were bisected and aggregated. The results obtained from the these experiments were summuerized as follows: 1. When mouse and rabbit 8-, 16-cell and morular embryos were culature in H-Y antiserum, the ratio of embryo which has developed to hatching blastocyst was 53.4, 46.3 and 57.4% in mouse embryos, and 49.0, 52.0 and 61.0% in rabbit embryo, respectively. The ratio of mouse and rabbit embryos developed to hatching blastocyst showed nearly natural sex rate(50%), except rabbit mourla showed a little higher ratio(61.0%) as compared to natural sex ratio. 2. When rabbit demi-embryos from 8-, 16-cell embryo and morula were cultured, the percentage of demi-embryos was 70, 68 and 58% without zona pellucida removed, and 62, 69 and 51% with zona pellucida. The rate of aggregation was higher in 8- and 16-cell demi-embryos than in morula demi-embryo. 4. When sexed-demi-embryo was aggregated with another demi-embryo with demi-embryo with same sex, the rate of embryo developed to blastocyst was 60, 50 and 25%, respectively. Eight-cell demi-embryo showed highest rate. In conclusion, it showed that H-Y antiserum which was made by rat spleen cell enabled sexing rabbit embryos. And when rabbit sexed 8-, 16-cell and morula demi-embryo were aggregated, they were developed to eu-blastocyst which suggested the potential of sexed embryo manipulation.

• Health Policy and Management
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• v.14 no.3
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• pp.97-121
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• 2004

Recently, human embryonic stem cell research raises exciting public expectation on medical possibilities as well as ethical debate. Embryo management has become an integral part of the management of infertility treatment, researches on embryo and human embryonic stem cells and so on. Britain has permitted the research on stem cells derived from human embryo which made the first nation to allow the cloning of human embryo for the stem cell research. However, new technologies such as the assisted reproductive technologies and human embryonic stem cell research continue to pose an increasing source of ethical dilemmas for physician, scientists, legislators, religious authorities and the general publics to deal with. None the less, the United Kingdom has adopted the most liberal policies regarding human embryo and human embryonic stem cell research. The implication of the British embryo management system are as follows: 1) the development of reproductive technologies and new stem cell research technologies continue to pose legal and ethical debates, since those involve several parties; 2) the UK has taken the legal and institutional approaches to cope with those serious issues; 3) the UK adopted most liberal policies regarding embryonic and human embryonic stem cell researches; 4) the British HFE Act is consistent with the existing Acts related to human embryo management and researches; 5) through amending the HFE Act to accomodate the changes of technologies, the UK try to minimize the legal and ethical burden on undertaking research regarding embryo. The debates about the researches on human embryo and human embryonic stem cells is likely to continue in the Korean society. Because of the controversy and competing ethical values, as well as the evolving technologies, so far no consensus exists in our society. It suggest that it is premature to bring closure by ruling out any particular approaches. Thus our society needs to make an efforts to find a basis which could resolve the societal controversies through enriching the societal conversation about the profound ethical issues regarding embryo management.

• Environmental Mutagens and Carcinogens
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• v.21 no.1
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• pp.44-50
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• 2001

To identify nongenotoxic carcinogen determined as negative by ICH guideline-recommended standard genotoxicity test battery; Ames test, chromosome aberration assay, mouse lymphoma $tk^{+/-}$ assay, in vivo micronucleus assay, we picked bisphenol A as a model compound. In this study, we applied in vitro BalB/c 3T3 cell transformation assay and Syrian hamster embryo (SHE) cell transfarmation assay. Bisphenol A was treated upto $769.2 ug/m{\ell}$ in BalB/c 3T3 cells and upto $125 ug/m{\ell}$ in SHE cells. bisphenol A didn't induced morphological transformation both with one stage treatment protocol and with two stage treatment protocol. But, treated far 48 hr, Bisphenol A induced morphological transformation significantly in SHE cells.

• Clinical and Experimental Reproductive Medicine
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• v.47 no.2
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• pp.114-121
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• 2020

Objective: Despite extensive research on implantation failure, little is known about the molecular mechanisms underlying the crosstalk between the embryo and the maternal endometrium, which is critical for successful pregnancy. Profilin 1 (PFN1), which is expressed both in the embryo and in the endometrial epithelium, acts as a potent regulator of actin polymerization and the cytoskeletal network. In this study, we identified the specific role of endometrial PFN1 during embryo implantation. Methods: Morphological alterations depending on the status of PFN1 expression were assessed in PFN1-depleted or control cells grown on Matrigel-coated cover glass. Day-5 mouse embryos were cocultured with Ishikawa cells. Comparisons of the rates of F-actin formation and embryo attachment were performed by measuring the stability of the attached embryo onto PFN1-depleted or control cells. Results: Depletion of PFN1 in endometrial epithelial cells induced a significant reduction in cell-cell adhesion displaying less formation of colonies and a more circular cell shape. Mouse embryos co-cultured with PFN1-depleted cells failed to form actin cytoskeletal networks, whereas more F-actin formation in the direction of surrounding PFN1-intact endometrial epithelial cells was detected. Furthermore, significantly lower embryo attachment stability was observed in PFN1-depleted cells than in control cells. This may have been due to reduced endometrial receptivity caused by impaired actin cytoskeletal networks associated with PFN1 deficiency. Conclusion: These observations definitively demonstrate an important role of PFN1 in mediating cell-cell adhesion during the initial stage of embryo implantation and suggest a potential therapeutic target or novel biomarker for patients suffering from implantation failure.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2002.11a
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• pp.73-73
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• 2002

Nuclear transfer (NT) techniques have advanced in the last years, and cloned animals have been produced by using somatic cells in several species including pig. However, it is difficult that the nuclear transfer porcine embryos development to blastocyst stage overcoming the cell block in vitro. Abnormal segregation of chromosomes in nuclear transferred embryos on genome activation stage bring about embryo degeneration, abnormal blastocyst, delayed and low embryo development. Thus, we are evaluated that the correlations of the frequency of embryo developmental rates and chromosome aberration in NT and In viかo fertilization (IVF) derived embryo. We are used for ear-skin-fibroblast cell in NT. If only karyotyping of embryonic cells are chromosomally abnormal, they may difficultly remain undetected. Then, we evaluate the chromosome aberrations, fluorescent in situ hybridization (FISH) with porcine chromosome 1 submetacentric specific DNA probe were excuted. In normal diploid cell nucleus, two hybridization signal was detected. In contrast, abnormal cell figured one or three over signals. The developmental rates of NT and IVF embryos were 55% vs 63%, 32% vs 33% and 13% vs 17% in 2 cell, 8 cell and blastocyst, respectively. When looking at the types of chromosome aberration, the detection of aneuploidy at Day 3 on the embryo culture. The percentage of chromosome aneuploidy of NT and IVF at 4-cell stage 40.0%, 31.3%, respectively. This result indicate that chromosomal abnormalities are associated with low developmental rate in porcine NT embryo. It is also suggest that abnormal porcine embryos produced by NT associated with lower implantation rate, increase abortion rate and production of abnormal fetuses.

• Plant Resources
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• v.5 no.2
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• pp.141-154
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• 2002

In this experiment, three kinds of mutations(ge, re, and eml )relating to the size of embryos were used to study their generation, genetic mechanism and developmental characteristics, and the interactions between embryo and endosperm were also examined. Giant embryo mutation comprises 7 kinds including the already isolated ge, and ge-2, which share an identical gene site. The SAM and the size of radicule for the ge showed little difference compared to a normal type. The number of embryo cells did not increased as much as it would affect the size of embryo. Therefore, the enlargement of embryo was due to the enlargement of scutellum that originated from the corpulence of each cell. Both F$_1$' s of re ]and odm 49 formed reduce embryos, and other combinations of hybridization showed all wild type of embryo sizes. Accordingly, the odm 49 must have an identical gene site of re 1, while odm 48 and odm 62 have different gene sites. Their shoots and radicules also shrank by the same ratio, however no sign of physical change was noticed. The size of embryo cell showed no change, while the number of cells was the half of that of wild types. The three gene sites of re represent all of them control the size of the entire embryo forming organs. The eml 1 was defined to have temperature sensibilities that the generation of endosperms was active at a high temperature while that was hampered at a low temperature.

• Clinical and Experimental Reproductive Medicine
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• v.28 no.1
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• pp.55-63
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• 2001

Objectives: This study was carried out to evaluate the effects of embryonic stage, cryoprotectant, and freezing-thawing method on the rates of survival and development of the cryopreserved mouse early embryo and finally to establish the cryopreservation method of surplus embryos obtained during assisted reproductive technology (ART). Materials and Methods: Two to eight cell embryos were obtained from oviducts of mated $F_1$ hybrid female mice superovulated by pregnant mare's serum gonadotropin (PMSG) and human chorionic gonadotropin (hCG). Two-step 1,2-propanediol (PROH), dimethylsulfoxide (DMSO) and 4-step PROH DMSO were used as cryoprotectant and dehydration and rehydration method of embryos, and slow-cooling or rapid-cooling method was used as frozen program. The survival rates of embryos were measured after thawing and rehydration, and the developmental rates of embryos were compared and observed during culturing embryos for 24, 48, 72, 96 hrs. Results: As for the survival and development rates of embryos according to embryonic stage, the survival rate of 2 cell stage in PROH and DMSO was significantly higher than 4-8 cell (64.5% versus 62.1 %,79.7% versus 73.2%) (p<0.01, p<0.01), but the development rates of 4-8 cell embryos in PROH and DMSO were significantly higher than 2 cell embryos for whole culture period (p<0.01) and the development rates of 4-8 cell embryos in PROH were significantly higher than 2 cell embryos in DMSO (p<0.01). As for the survival and development rates of embryos according to cryoprotectant, the survival rate of 2 cell embryo in DMSO was significantly higher than that in PROH (74.4% versus 64.5%) (p<0.01), whereas the development rate of embryos was not differ till 24 hrs. The developmen1 rate from morular to hatching blastocyst, however, was significantly higher in PROH than in DMSO during 48 hr (p<0.01). The survival rate of 4-8 cell embryo was 62.1% in PROH and 73.2% in DMSO. The development rates of embryo in PROH were significantly higher for whole culture periods (p<0.01, 0.05). In respect to the effect of freezing and thawing program on the survival and development rates of embryos, method of slow cooling and rapid thawing was more effective than that of rapid cooling and rapid thawing. Conclusions: The survival rate of embryo in 2 cell stage was higher than in 4-8 cell stage, and PROH appears more effective cryoprotectant than DMSO because PROH showed better development rates of embryos in 2 and 4-8 cell stage. Moreover, slow cooling and rapid thawing method was considered as the best cryopreservation program.

• Journal of Embryo Transfer
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• v.28 no.2
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• pp.141-147
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• 2013

The purpose of this study was attempted to new methods in mammalian embryos vitrification. This method was affected to increase of the embryo vitrification efficiency and it would be applied to the field of embryo transfer to recipient by modified loading method of embryo into 0.25 ml plastic straw. The frozen mouse embryos were carried out warmed from two different cell stages (8-cell and blastocyst, respectively) by attachment of an embryo in the vitrification straw (aV) method. All groups were cultured in M-16 medium to determine the development and survivability for 24 h, respectively. Results shown that, the survivability of two different groups were significantly different (94.8% vs. 70.9%). Total cell number was not significantly different the non-frozen blastocyst ($99.7{\pm}12.4$) compared to the post-thaw blastocyst ($94.8{\pm}15.1$). From the 8-cell embryo, total cell number of frozen blastocysts were significantly lower than others groups ($74.7{\pm}14.6$, p<0.05). In the case of cell death analysis, the blastocysts from non-frozen and frozen-thawed 8-cell group were not different ($0.0{\pm}0.0$ vs. $1.9{\pm}3.1$, p>0.05). However, the apoptotic nuclei of blastocyst were significantly observed the frozen-thawed group ($5.4{\pm}4.4$) compared to non-frozen group (p<0.05). Therefore, this new method of embryos using in-straw dilution and direct transfer into other species would be more simple procedure of embryo transfer rather than step-wise dilution method and cryopreservation vessels, so we can be applied in animal as well as human embryo cryopreservation in further.

• Journal of Genetic Medicine
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• v.18 no.1
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• pp.1-7
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• 2021

The oocyte quality is of great importance in infertility as it reflects the follicle developmental potential and further affects the embryo development, clinical pregnancy outcomes. The analysis of gene expression in somatic cells is an important study to better clinical in vitro fertilization (IVF) outcomes in embryo selection reflecting the appropriate communication between the oocyte and somatic cells. Specifically, somatic cell transcriptomic technology can help assess biomarkers of oocyte and embryo ability. The present article aims to overview the basic aspect of folliculogenesis and review studies involving changes in candidate gene expression of cumulus or granulosa cell related to clinical outcomes in human IVF.