• Journal of Embryo Transfer
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• v.10 no.1
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• pp.53-63
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• 1995

The effects of different protein sources (serum vs bovine serum albumin), growth factors (EGF and PDGF) and co-culture with various type of somatic cel1s (BOEC, MEF and BRL) on the in vitro development of in vitro matured / in vitro fertilized bovine oocytes were examined, and the viability of frozen/thawed embryos derived from IVM /IVF was examined. Cell numbers of blastocysts were also counted. In Experiment 1, CR$_1$aa with serum was superior to CR$_1$aa with BSA in producing morulae plus blastocysts from IVM /IVF oocytes(24.4% vs 30.4%, p>0.05). In Experiment 2, more morulae plus blastocysts(42.3%) were produced in CR$_1$aa containing long /ml EGF than in the control CR$_1$aa(33.3%). In Experiment 3, 2- to 8-cell embryos derived from IVM /IVF oocytes were randomly allotted to one of 4 culture groups : a) CR$_1$aa ; b) CR$_1$aa + ing /ml PDGF ; CR$_1$aa + Sng /ml PDGF ; CR$_1$aa + lOng /ml PDGF ; culture resulted in 21.3, 51.2, 41.4 and 45.9%(p<0.05), respectively, developing into morulae and blastocysts. In Experiment 4, 0 and Sng /ml PDGF added to CR$_1$aa coculture with BRL or BOEC yielded 47.5, 42.5, 33.8 and 41.6% morulae and blastocysts, respectively. In Experiment 5, the proportion of embryos into morulae and blastocysts was highest in CR$_1$aa with MEF coculture group(50.9%) compared to any other group(CR$_1$aa, 22.3%; CR$_1$aa+BRL, 32.9%; CR$_1$aa+BOEC, 33.8%, p>0.05). In Experiment 6, survival rate of blastocysts produced by in vitro fertilization when cryoprotectant was removed in 0.7M glycerol+0.7M sucrose and 0.7M sucrose solution for 10 min. after thawing at 2$0^{\circ}C$ (Exp. H, 58.8%) was slightly higher than when cryoprotectant was removed 10%, 6.7% and 3.3% glycerol for 10 min. after thawing at 37$^{\circ}C$ (Exp. I, 54.3%). These study indicate that growth factors and somatic cell co-culture can increase the proportion of embryos that develop into morulae and blastocysts without an increase in the cell number and frozen /thawed method employed this experiment was not different.

• Journal of Embryo Transfer
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• v.21 no.2
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• pp.121-128
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• 2006

In this study, we examined the effects of molecular weight, concentrations and treat the duration of polyvinylpyrrolidone (PVP) in vitro maturation (IVM) medium (Experiment 1), and the effect of PVP in IVM, in vitro fertilization (IVF) and in vitro culture (IVC) medium on the development and cell number of porcine embryos (Experiment 2). The base mediums were NCSU 23 solution for IVM, mTBM solution for IVF and PZM3 solution for IVC. In experiment 1, the development rates to 2 cell and blastocyst stage were not differ from the different molecular weight (MW), concentration and duration of PVP in IVM medium. However, the hatching rate of blastocyst was significantly higher in the group of MW 40,000, 0.5% and $0{\sim}44hr$ than in the other groups (p<0.05). In experiment 2, the results of IVM, IVF and IVC medium with (W) or without (W/O) 0.5% MW 40,000 PVP are follows. The development rate to 2 cell stage was highest in the group of W-W/O-W (p<0.05). The development rate to blastocyst and hatching rate was higher in the group of W-W/O-W and W-W/O-W/O than that of other treatments (p<0.05).

• Journal of Embryo Transfer
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• v.13 no.1
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• pp.77-86
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• 1998

A combined technology of transvaginal ovum pick-up(OPU) system with in vitro-oocyte manipulation technique can be used for improving reproductive efficiency in the cattle. The objective of this study was to establish a newly-conceived breeding program using OPU in the pregnant cows. The OPU trial was performed in pregnant cows every 10 days from 40 through 90 days of artificial insemination (Al), and number of follicles in ovary, number of retrieved oocytes and embryo development following in vitro-fertilization, were evaluated. Reduced number of follicles in the ovaries of pregnant cows was firstly detected from 70 days after A' and a significant (P<0.05) decrease in the follicle number (5.4 follicles /donor) was found at 90 days than at 40, 50, 60 and 80 days after Al (8.0~9.2). A similar pattern was also observed in the number of oocytes retrieved by OPU apparatus during experimental period. When retrieved oocytes were matured and inseminated in vitro with frozen bull semen, development of the oocytes to the blastocyst stage was not significantly affected by the retrieval time. Four embryos (morula or blastocyst stage) derived from oocytes retrieved from pregnant cows were nonsurgically transferred to four recipient cows on day 7 of estrus cycle. For the first time in Korea, three of four transferred embryos developed to live calves with normal physiological parameters. In conclusion, an effective breeding program employing pregnant cow can be developed by use of OPU trial and in vitro culture techniques of oocytes ; OPU system could be repeated in pregnant cows with no risk of abortion and viable offsprings were borne after transfer to the recipients.

• Korean Journal of Animal Reproduction
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• v.20 no.2
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• pp.163-170
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• 1996

The effect of several potential antioxidants and amino acids were examined as a means of increasing the development of in vitro matured and in vitro fertilized oocytes into morulae or blastocysts. Bovine embryos developed to the 2~8 cell stage after in vitro fertilization were cultured for 5 to 6 days at 39$^{\circ}C$ in CR1aa containing varing concentraton of the antioxidants and amino acid in a gas phase consisting of 5% CO2, high humidified air. At 5~6 days, embryo developments were reduced, and embryos were fixed and stained with Hochest 33342 DNA stain to facilitate counting of cells. In experiment 1, the proportion of embryos developed to morulae and blastocysts in CR1aa containing 1mM, 2.5mM taurine (22.6% and 20.4%) was slightly higher than those of 0, 5 and 10mM Taurine (5.7, 5.7 and 3.9%, P<0.05). In experiment 2, addition of glutathione did not improve blastocyst development (P>0.05). In experiment 3, concentations of superoxide dismutase(SOD) ranging from 300 to 1,000 U did not affect the propotion of embryos developing into blastocysts (P>0.05). In experiment 4, addition of 250 U catalase(38.5%) was slighty higher than those of 0, 500 and 1,000U. In experiment 5, the proportion of embryo developed beyond morula stage in CR1aa with taurine plus EDTA was slighty higher than other treatments(15.7, 26.0 and 29.2%), there were no significantly increases in cell number among treatments(P>0.05). These results are indicating that antioxidants and amino acids can increase the proportion of embryos that develop into morulae and blastocysts, but did not increas in cell number of blastocysts.

• Clinical and Experimental Reproductive Medicine
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• v.42 no.2
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• pp.45-50
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• 2015

Objective: Artificial oocyte activation (AOA) is an effective method to avoid total fertilization failure in human in vitro fertilization-embryo transfer (IVF-ET) cycles. AOA performed using a calcium ionophore can induce calcium oscillation in oocytes and initiate the fertilization process. We evaluated the usefulness of AOA with a calcium ionophore in cases of total fertilization failure in previous cycles and in cases of severe male factor infertility patients with non-motile spermatozoa after pentoxifylline (PF) treatment. Methods: The present study describes 29 intracytoplasmic sperm injection (ICSI)-AOA cycles involving male factor infertility at Cheil General Hospital from January 2006 to June 2013. Patients were divided into two groups (control, n=480; AOA, n=29) depending on whether or not AOA using a calcium ionophore (A23187) was performed after testicular sperm extraction-ICSI (TESE-ICSI). The AOA group was further split into subgroups according to sperm motility after PF treatment: i.e., motile sperm-injected (n=12) and non-motile sperm-injected (n=17) groups (total n=29 cycles). Results: The good embryo rate (52.3% vs. 66.9%), pregnancy rate (20.7% vs. 52.1%), and delivery rate (10.3% vs. 40.8%) were lower in the PF/AOA group than in the control group. When evaluating the effects of restoration of sperm motility after PF treatment on clinical outcomes there was no difference in fertilization rate (66.6% vs. 64.7% in non-motile and motile sperm, respectively), pregnancy rate (17.6% vs. 33.3%), or delivery rate (5.9% vs. 16.7%) between the two groups. Conclusion: We suggest that oocyte activation is a useful method to ensure fertilization in TESE-ICSI cycles regardless of restoration of sperm motility after PF treatment. AOA may be useful in selected patients who have a low fertilization rate or total fertilization failure.

• Clinical and Experimental Reproductive Medicine
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• v.25 no.2
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• pp.123-128
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• 1998

Implantation rates remain low following human in vitro fertilization (IVF). Suboptimal culture conditions may limit the ability of embryos to hatch as blastocysts, and artificial opening of the zona pellucida has been proposed as a means to promote subsequent hatching (assisted hatching). In this study, assisted hatching (AH) by zona drilling using acidic Tyrode's solution was performed in 320 patients, due to their age of more than 38 years (group A), the thick zona pellucida (group Z; $ZP\geq0.18{\mu}m$), and failures in implantation more than 3 times in previous IVF-ET trial (group P). This study was designed firstly, to study the effects of AH on the outcomes of IVF-ET according to the indications and secondly, to verify the appropriate application of AH. The results were as follows; 1. There was no difference in pregnancy rate between AH group (26.6%) and non-AH group (26.5%). 2. Assisted hatching (AH) showed significantly higher pregnancy rate of the patients with thick zona pellucid a than those of the patients with age factor and with the history of repeated implantation failure. But in the patients with age factor only, AH resulted in higher pregnancy rate. 3. Interestingly, the patients with complex factors including zona factor (Z: 33.9%; ZA: 30.4%; ZP: 31.6%; ZAP: 21.4%) showed higher pregnancy rates than other complex factors excluding zona factor (A: 24.4%, P: 0%; AP: 10.8%). From these results, AH is more helpful to the patients with thick zona pellucida rather than patients with older age and/or previous repeated implantation failure.

• Neonatal Medicine
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• v.18 no.2
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• pp.197-203
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• 2011

Purpose: The purpose of this study is to compare perinatal outcomes between in vitro fertilization (IVF) twins and naturally conceived twins born to women aged 35 years or older and to provide basic information for taking care of IVF twins born to women aged 35 years or older. Methods: We reviewed the records of perinatal and neonatal outcomes in 288 IVF twins and 220 naturally conceived twins born to women aged 35 years or older between January 2001 and December 2010 at CHA Bundang Medical Center. Results: No difference was observed in the maternal ages of mothers giving birth to IVF twins and those giving birth to naturally conceived twins. Gestational ages and birth weights of IVF twins were not different from those of naturally conceived twins. Various perinatal outcomes, including gestational diabetes mellitus, pregnancy-induced hypertension, placenta previa, premature amniotic membrane rupture, and need for a Cesarean section did not differ between the 2 groups. However, the 1-min and 5-min Apgar scores (P=0.019 and P=0.045, respectively) were different between the 2 groups. The incidence of early-onset sepsis was lower in the IVF twins than in the naturally conceived twins (P=0.02). However, the 2 groups did not show any difference in the incidence of respiratory distress syndrome, bronchopulmonary dysplasia, patent ductus arteriosus, necrotizing enterocolitis, intraventricular hemorrhage, and other congenital anomalies. Conclusion: The perinatal outcomes in IVF twins born to women aged 35 years or older were not significantly different from those of naturally conceived twins.

• Journal of Embryo Transfer
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• v.18 no.3
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• pp.237-242
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• 2003

These studies were carried out to establish an effective in vitro embryo transfer methods by analyzing several factors. The base media were TCM-199 solution for in vitro maturation(IVM) of bovine follicular oocytes and Fer-TALP solution for in vitro fertilization(IVF) and CRlaa medium for in vitro culture(IVC). IVC used the fertilized oocytes of 24-hr culture (day 1)after IVF. Embryos were cultured in drop-culture that contained 25 embryos per 10${mu}ell$. Blastocysts cultured for 7 to 9 in vitro were transferred to recipients. The results obtained were as follows: 1. The pregnancy rate according to different region of embryo transfer were 33.8％, 48.1％, 45.0％ and 35.3％ respectively. 2. The pregnancy rate according to the parity of recipient when embryos were transferred to nulliparous (42.9％) was higher than that of 1∼3nd parlous(36.9％), however there were not show significant difference each other. 3. According to the stage of blastocyst, the pregnancy rate when middle blastocysts (MB) (45.5％) were transferred to recipients were higher than that of late blastocysts (LB) (41.0％). 4. When IVF-derived blastocysts cultured for 7 to 9 day were transferred to recipients, the pregnancy rate was higher 7 day of blastocyst than that of 8 day or 9 day of blastocyst. The results of embryo transfer according to the regions, the parity of recipient and the development stage showed that blastocyst formed for 7day transferred to nulliparou were higher pregnancy rate than others.

• Korean Journal of Animal Reproduction
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• v.24 no.3
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• pp.311-317
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• 2000

This study was carried out to compare the temperature and time of heat shock, and the effect of heat shock on development of embryos after in vitro maturation and fertilization in bovine oocytes. The results obtained were as follows. 1. The optimum temperature and time of heat shock were 41$^{\circ}C$ and 30sec on in vitro development of embryos from 4~8 cell to blastocyst. 2. The rates of cleavage on zygotes produced on in vitro were significantly increased by heat shock after IVM than before IVM(P<0.05). 3. When the oocytes were treated heat shock after IVM and 5 days cultured, developmental rates to blastocyst were increased than other experimental treatments.

• Clinical and Experimental Reproductive Medicine
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• v.48 no.4
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• pp.352-361
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• 2021

Objective: The study assessed the developmental potential of germinal vesicle (GV) oocytes subjected to in vitro maturation (IVM) after prematuration culture with cilostamide (a phosphodiesterase-3 inhibitor) and the impact of cilostamide exposure on the morphology of meiosis II (MII) oocytes and subsequent embryo quality. Methods: In total, 994 oocytes were collected from 63 patients. Among 307 GV oocytes, 140 oocytes were selected for the experimental group and 130 oocytes for the control group. The denuded GV-stage oocytes were cultured for 6 hours with cilostamide in the experimental group and without cilostamide in the control group. After 6 hours, the oocytes in the experimental group were washed and transferred to fresh IVM medium. The maturational status of the oocytes in both groups was examined at 26, 36, and 48 hours. Fertilization was assessed at 18 hours post-intracytoplasmic sperm injection. Embryo quality was assessed on days 3 and 5. Results: In total, 92.1% of the oocytes remained in the GV stage, while 6.4% converted to the MI stage (p<0.01) after cilostamide exposure. In both groups, more MII oocytes were observed at 36 hours (25.8% vs. 21.5%) than at 26 hours (10.8% vs. 14.6%) and 48 hours (13% vs. 7.9%) (p>0.05). With the advent of cilostamide, blastocyst quality was better in the experimental group than in the control group (p<0.05). Conclusion: Cilostamide effectively blocked nuclear maturation and promoted cytoplasmic growth. Prematuration culture with cilostamide enabled synchronization between cytoplasmic and nuclear maturity, resulting in better blastocyst outcomes.