• Clinical and Experimental Reproductive Medicine
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• v.42 no.1
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• pp.14-21
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• 2015

Objective: It has previously been suggested that embryos developing from intracytoplasmic sperm-injected (ICSI) zygotes with three pronuclei (3PN) are endowed with a mechanism for self-correction of triploidy to diploidy. 3PN are also observed in zygotes after conventional in vitro fertilization (IVF). The parental origin, however, differs between the two fertilization methods. Whereas the vast majority of 3PN IVF zygotes are of dispermic origin and thus more likely to have two centrioles, the 3PN ICSI zygotes are digynic in origin and therefore, more likely to have one centriole. In the present study, we examine whether the parental origin of 3PN embryos correlates with the karyotype. Methods: The karyotype of each nucleus was estimated using four sequential fluorescence in situ hybridizations-each with two probes-resulting in quantitative information of 8 different chromosomes. The karyotypes were then compared and correlated to the parental origin. Results: 3PN ICSI embryos displayed a significantly larger and more coordinated reduction from the assumed initial 3 sets of chromosomes than 3PN IVF embryos. Conclusion: The differences in the parental origin-and hence the number of centrioles-between the 3PN IVF and the 3PN ICSI zygotes are likely to be the cause of the differences in karyotypes.

• Clinical and Experimental Reproductive Medicine
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• v.25 no.2
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• pp.129-134
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• 1998

The objective of this study was to test whether the developmental capacity of human multi-pronuclear (PN) zygotes after ultra-rapid freezing using EM grid can be maintained. For this experiment, multi-PN zygotes which produced in human IVF program were used as an alternatives of normal 2PN zygotes, and they were separated into 3PN or $\geq4PN$ zygotes to compare their in vitro development and cryoinjury according to PN number. As freezing solution, EFS30 which consisted of 30% ethylene glycol, 18% bcoll, 0.5 M sucrose and 10% FBS added D-PBS was used. The result obtained in this experiment was summarized as follows; When the multi..PN zygotes were ultrarapidly frozen and thawed, the high mean percentages (85.5%) were survived. Also when the cleavage rates between control and freezing group were compared with PN number, there were not significantly different in each group (3PN; 81.3% & 85.4% and $\geq4PN$; 90.0% & 95.7%). When the in vitro development rates after thawing were examined, freezing 3PN group (22.0%) was not differed to control 3PN group (38.5%), although the development result of freezing $\geq4PN$ group (45%) was significantly lower than that of control $\geq4PN$ group (44.4%) (p<0.05). These results demonstrate that developmental capacity of human zygote can be obtained by ultra-rapid freezing method using EM grid and EFS30.

• Clinical and Experimental Reproductive Medicine
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• v.37 no.4
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• pp.329-338
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• 2010

Objective: This study was carried out to know whether cryopreservation of sibling 2PN zygotes could increase the cumulative delivery rates in the patients who had less than 10 fertilized zygotes. Methods: A retrospective analysis was performed in 138 in vitro fertilization-embryo transfer (IVF-ET) cycles with less than 10 fertilized zygotes during January 2003 to December 2007 in Cheil General Hospital. These cycles were divided into two groups. In Group I (n=86), all fertilized embryos were cultured to transfer on day 3 without cryopreserved embryos at the 2PN stage. In Group II (n=52), among fertilized zygotes, some sibling zygotes were frozen at the 2PN stage, the remainder were cultured to transfer. Clinical outcomes in fresh ET cycles and cumulative ongoing pregnancy rates after subsequent frozen-thawed (FT)-ET cycles were compared. Results: There were no significant differences in female mean age, number of retrieved oocytes and total fertilized embryos between two groups, Number of cultured embryos was significantly lower in Group II ($5.2{\pm}0.5$) than in Group I ($8.4{\pm}0.7$) (p<0.01). Also, number of transferred embryos was significantly lower in Group II ($3.3{\pm}0.6$) compared with Group I ($3.6{\pm}0.6$) (p<0.01). ${\beta}$-hCG positive rates and delivery rates (51.2 vs. 46.2 % and 41.9 vs. 34.6 %, respectively) after fresh ET were slightly higher in Group I than in Group II. However, the differences were not statistically significant. Also, the cumulative delivery rates after subsequent FT-ET cycles were not significantly different between Group I (48.8%) and Group II (50.0%). Conclusion: This study showed that cryopreservation of sibling 2PN zygotes from patients who had less than 10 zygotes in the fresh ET cycles did not increase cumulative delivery outcomes. But, it could provide an alternative choice for patients due to offering more chance for embryo transfers if pregnancy was failed in fresh IVF-ET cycles.

• Clinical and Experimental Reproductive Medicine
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• v.50 no.4
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• pp.244-252
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• 2023

Objective: We evaluated the efficacy of the newly developed optimized in vitro culture (OIVC) dish for cultivating preimplantation mouse embryos. This dish minimizes the need for mineral oil and incorporates microwells, providing a stable culture environment and enabling independent monitoring of individual embryos. Methods: Mouse pronuclear (PN) zygotes and two-cell-stage embryos were collected at 18 and 46 hours after human chorionic gonadotropin injection, respectively. These were cultured for 120 hours using potassium simplex optimized medium (KSOM) to reach the blastocyst stage. The embryos were randomly allocated into three groups, each cultured in one of three dishes: a 60-mm culture dish, a microdrop dish, and an OIVC dish that we developed. Results: The OIVC dish effectively maintained the osmolarity of the KSOM culture medium over a 5-day period using only 2 mL of mineral oil. This contrasts with the significant osmolarity increase observed in the 60-mm culture dish. Additionally, the OIVC dish exhibited higher blastulation rates from two-cell embryos (100%) relative to the other dish types. Moreover, blastocysts derived from both PN zygotes and two-cell embryos in the OIVC dish group demonstrated significantly elevated mean cell numbers. Conclusion: Use of the OIVC dish markedly increased the number of cells in blastocysts derived from the in vitro culture of preimplantation mouse embryos. The capacity of this dish to maintain medium osmolarity with minimal mineral oil usage represents a breakthrough that may advance embryo culture techniques for various mammals, including human in vitro fertilization and embryo transfer programs.

• Journal of The Korean Society of Grassland and Forage Science
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• v.38 no.4
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• pp.320-324
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• 2018

In this study, we examined effect of sperm penetration of oocytes after in vitro fertilization (IVF) with cauda epididymal spermatozoa in Hanwoo bull after feeding of timothy hay. One testicle with epididymides was castrated from one Hanwoo bull (14 months of age) and spermatozoa recovered from cauda epididymis and cryopreserved. As control, frozen Hanwoo semen was used. Matured cumulus oocyte complexes were co-incubated with frozen-thawed cauda epididymal spermatozoa for 12 or 18 hours. After IVF, presumptive zygotes were cultured in modified synthetic oviductal fluid. In experiment 1, we examined sperm penetration rate at 12 hours of IVF with epididymal sperm. Total penetration rate among cauda epididymis and control was similar(mean${\pm}$standard error, cauda epididymis and control vs. $49.7{\pm}11.3$ and $54.4{\pm}12.8%$). In experiment 2, cleavage and blastocyst developmental rate were evaluated at day 2 and day 8 after IVF for 18 hours. Cleavage rate among cauda epididymis and control was similar(cauda epididymis and control vs. $81.2{\pm}3.4$ and $82.7{\pm}2.5%$). However, blastocyst developmental rate of cauda epididymis group was significantly higher than that of control group(cauda epididymis and control vs. $24.4{\pm}1.6$ and $12.2{\pm}2.8%$, p<0.05). In conclusion, cauda epididymal spermatozoa in Hanwoo bull has high embryo developmental competence and can be used as an alternative to ejaculated frozen sperm in vitro.

• Clinical and Experimental Reproductive Medicine
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• v.27 no.1
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• pp.59-65
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• 2000

Objective: The purpose of this study was to determine the important factors affecting survival and pregnancy rate in frozen-thawed embryo transfer cycles. Methods: we performed retrospective analysis in 738 cycles of frozen-thawed embryo transfers, in relation to the insemination methods, the freezing stage of embryo, patient's age, infertility factors and the origin of injected sperm in ICSI cycles. After conventional IVF or ICSI, the supernumerary PN stage zygotes or multicellular embryos were cryopreserved by slow freezing protocol with 1,2-propanediol (PROH) as a cryoprotectant. Results: The survival rates of thawed embryos were 69.3% (1585/2287) in conventional IVF group and 71.7% (1645/2295) in ICSI group. After frozen-thawed embryo transfers, 27.0% (92/341) and 32.0% (109/341) of pregnancy rates were achieved in conventional IVF and ICSI group, respectively. There were no significant difference in the survival and pregnancy rates according to the insemination methods, the freezing stage and patient's age. However, the pregnancy rate (36.2%) of male factor infertility was significantly higher than the tubal (27.2%) and other female factor infertility (22.9%). In ICSI group, the origin of injected sperm did not affect the outcome of frozen-thawed embryo transfer cycles. Conclusion: The present study demonstrates that acceptable clinical outcomes can be achieved after the transfer of frozen-thawed embryos regardless of the stage of embryos for freezing, the patient's age and the origin of injected sperm.