• Clinical and Experimental Reproductive Medicine
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• v.28 no.4
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• pp.287-293
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• 2001

Objective : To investigate the efficacy of high infusion frequency of liquid nitrogen on pregnancy in human embryo after freezing and thawing. Materials and Methods: 150 infertile patients underwent 162 consecutive thawing-ET cycles. In the high infusion frequency group (Group A), 47 patients (50 cycles) underwent cryopreservation with high infusion frequency of liquid nitrogen. In the low infusion frequency group (Group B), 103 patients (112 cycles) underwent cryopreservation with low infusion frequency of liquid nitrogen. We analyzed the clinical characteristics, fertilization rates, development of embryo, good quality embryo ratio, implantation rates, and pregnancy rates between these two groups. Results: There was no difference between the groups with regard to clinical characteristics (mean age, infertility duration, infertility factors, hormone profile), mean number of oocyte retrieval, fertilization rates, and mean embryo number of transfers. The survival rates in group A was 64.9% (228 of 350 embryos), and among the 228 embryos 190 embryos (83.3%) which progressed to the two- to eight-cell stage. After thawing, the embryo numbers were 65 (34.2%), 29 (15.3%), 35 (18.4%), and 37 (19.5%) of grades 1, 2, 3, and above 4, respectively. The survival rates in group B was 63.8% (482 of 755 embryos), and among the 482 embryos 465 embryos (96.5%) which progressed to the two- to eight-cell stage. After thawing, the embryo numbers were 106 (22.8%), 94 (20.2%), 89 (19.1%), and 112 (24.1%) of grades 1, 2, 3, and above 4, respectively. There was no difference in embryo quality change after the freezing-thawing procedure between the groups. Implantation rates (31.1% vs. 34.3%) were not significant. However hCG positive rates in group A (40%) were higher than group B, but not statistically significant. Clinical pregnancy rate (26% vs. 25.9%), on going pregnancy rates (>20 weeks) were not significant (26% vs. 25%). Conclusion: We compared embryo quality change, survival rates, and pregnancy rates between high infusion frequency group and low infusion frequency group and the results were similar between the two groups. Therefore, high infusion frequency of liquid nitrogen for cryopreservation is a worthy method to preserve in human embryos.

• Asian-Australasian Journal of Animal Sciences
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• v.20 no.1
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• pp.31-35
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• 2007

The main purpose of this study was to improve the efficiency and quality of in vitro embryo production in Korean Native Cows (KNC). We examined the effects of ovarian morphologies (Experiment 1) and the culture vessel (Experiment 2) on in vitro maturation (IVM). We measured the subsequent development rates and cell numbers of blastocysts. In Experiment 1, the ovaries of KNC were divided into six groups, based on follicle and corpus luteum (CL) morphology. The development rates to the 2- and 8-cell stages were similar among the six groups. The development rates to blastocyst stages were significantly higher in the group without a CL or follicle (WOCL/F) than in the groups with follicular cysts (FCs), regressive CLs (RCLs) or cystic CLs (CCLs) (p<0.05). The cell number of the inner cell mass (ICM) of blastocysts in the FCs and RCLs groups, and the number of cells in the trophectoderm (TE) in the WOCL/F group, FCs, growing CLs (GCLs) and RCLs were significantly higher than in other groups (p<0.05). The total cell number (TCN) in the WOCL/F, FC and RCL groups was also significantly higher than in other groups (p<0.05). The ICM cell number/TCN ratio was significantly higher in the FC and RCL groups than in the GCL and DF groups (p<0.05). In Experiment 2, oocyte IVM was carried out in culture dishes, in 0.25- or 0.5-ml straws used for freezing sperm. The development rate to the 2-cell stage was significantly higher in the 0.5-ml straw group than in the 0.25-ml straw group. The development rates to the blastocyst stage were similar in the dish and the two straw groups. There were no differences in the cell numbers of ICM, TE or TCN or ICM cell number/TCN ratios between groups.

• Development and Reproduction
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• v.1 no.1
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• pp.67-77
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• 1997

Mammalian ovary consists of various growing stages of follicles. Ovarian follicular growth and differentiation, however, can be distinguished into recruitment, growth, selectiona nd ovulation. while only minute of the selected follicles ovulate their oocytes, all the rest follicles disappear by atresia. this atresia is an important event of which physiological mechanism must be resolved. The present study was carried out to investigate the effects of various doses of pregnant mare's serum gonadotropin (PMSG) on the oocyte quality, ovulation rate, and the early embryonic development in immature mice. Immature mice were administrated with 5, 20, or 40 IU PMSG. At every 12 hour up to 72 hour after treatment, body and ovary weights were measured. Oocytes were flushed from the oviducts under the dissecting microscope and observed under the inverted microscope. Late 2-cell embryos were collected from the mice which were superovulated by the same dosage of PMSG followed by 5 IU hCG 47 hours after PMSG-treatment. The percentage of abnormal oocytes was higher in 20 or 40 IU PMSG-treated animals than 5 IU PMSG-treated ones. Ovulation occured at 12 hours afger PMSG injection in all experimental groups. The percentage of retrieved abnormal oocytes increased in the 20 or 40 IU PMSG-treated goups but not in 5 IU PMSG-treated group. There was no significant difference in the mating rate among the groups [52.6% (10/19), 66.7% (10/15), 44.0% (11/25) : 5, 20, 40 IU group respectively] ; however, ther was a significant (p<0.01) increase of embryo retrieval rates in 5 and 20 IU-treated groups compared with that in 40 IU-treated group [89.2% (239-268), 85.5% (224/262), 40.0% (18/45)]. There was significant (p<0.01) increase of embryo development rates in 5 IU-treated group compared with that in 20 and 40 IU-treated group [231/239(96.7), 179/224(79.9), 77.8(14/18)]. In conclusion, higher doses of PMSG injection increased the occurrence of abnormal oocytes ovulation in immature mice. The most of oocytes collected from 5 or 20 IU-PMSG-treated group has fertilizabioity. But in mice injected iwth higher doses of PMSG, their oocytes exhibit less fertilizability and, even fertilized, all oocytes are not fully capable of development.

• Clinical and Experimental Reproductive Medicine
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• v.46 no.4
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• pp.178-188
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• 2019

Objective: To determine the clinical pregnancy (CP) and live birth (LB) rates arising from frozen embryo transfers (FETs) that had been generated under the influence of in vitro fertilization (IVF) adjuvants given to women categorized as poor-prognosis. Methods: A registered, single-center, retrospective study. A total of 1,119 patients with first FETs cycle include 310 patients with poor prognosis (109 treated with growth hormone [GH], (+)GH group vs. 201 treated with dehydroepiandrosterone, (-)GH group) and 809 patients with good prognosis (as control, (-)Adj (Good) group). Results: The poor-prognosis women were significantly older, with a lower ovarian reserve than the (-)Adj (Good) group, and demonstrated lower chances of CP (p< 0.005) and LB (p< 0.005). After adjusting for confounders, the chances of both CP and LB in the (+)GH group were not significantly different from those in the (-)Adj (Good) group, indicating that the poor-prognosis patients given GH had similar outcomes to those with a good prognosis. Furthermore, the likelihood of LB was significantly higher for poor-prognosis women given GH than for those who did not receive GH (p< 0.028). This was further confirmed in age-matched analyses. Conclusion: The embryos cryopreserved from fresh IVF cycles in which adjuvant GH had been administered to women classified as poor-prognosis showed a significant 2.7-fold higher LB rate in subsequent FET cycles than a matched poor-prognosis group. The women with a poor prognosis who were treated with GH had LB outcomes equivalent to those with a good prognosis. We therefore postulate that GH improves some aspect of oocyte quality that confers improved competency for implantation.

• Fisheries and Aquatic Sciences
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• v.12 no.4
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• pp.276-285
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• 2009

Despite great commercial interest, relatively little has been described about molecular mechanism of bivalve reproduction. We investigated genes involved in ovarian maturation of the Yesso scallop, Patinopecten yessoensis. GSI index and histological analysis revealed that maturation of ovary begin in February and spawning period is from April to June which is similar to the previous study in the East Sea. As result of combination analysis of differential display RTPCR (DDRT-PCR) and histological examination, vitellogenin (Vg), ferritin (Ft) and ADT/ATP carrier protein (ACC) were identified as differently expressed genes in maturating ovary. Endpoint RT-PCR results showed that Vg is ovary-specific genes whereas Ft and ACC are expressed ubiquitously suggesting that Vg can be good molecular markers for ovarian development and sex determination in bivalves. Quantitative PCR results revealed that Vg were expressed highest during growth stage and appears to play a major role in oocyte maturation. On the contrary, expression of Ft was highest after spawning stage, which suggests that up-regulation may be involved in spawning and inactive stages in which the scallops recover from spawning. In addition, high level of the mitochondrial gene, ACC, may play a role in energy metabolism in maturating oocytes. Isolation and molecular studies of these key genes will expand our knowledge of the physiological changes from various exogenous factors including temperature, salinity, pH, even or numerous endocrine disrupting chemicals (EDCs) during reproductive cycle. In addition, further study of these genes implicates various industrial applications including the stable seed production, increased food quality, or economic aquaculture system.

• Proceedings of the Korean Society of Embryo Transfer Conference
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• 2002.11a
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• pp.79-79
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• 2002

본 연구에서는 in vitro에서 성숙된 난자의 핵성숙(Polar Body extrusion)에 소요되는 시간과 배반포 단계로의 발달능력 사이의 관계를 비교하여 조기에 발달능력을 가진 embryo를 선발할 수 있는 IVP 체계를 개발하고자 하였으며 in vitro maturation(IVM)에 따른 first polar body(PB) 형성, IVM과 IVF 시간이 oocyte의 발달에 미치는 영향과 생산된 배반포의 세포수를 평가하였다. IVM은 TCM199 배양액을 사용하였고 in vitro fertilization(IVF)은 Fer -TALP용액을 사용하였으며 in vitro culture(IVC)는 CRlaa 배양액을 사용하여 2일까지는 0.3% BSA를 3일 부터는 10%FBS와 bovine oviduct epithelial cell을 첨가하여 배양하였다. IVM 시간에 따른 PB의 출현율은 0hr(0%), 6hr(0%), 12hr(0%), 14hr(8.7%), 16hr(40.5%), 18hr(48.0%), 20hr(65%), 22(68%) 그리고 24hr(74.5%)을 보였으며 IVM 시간에 따른 cleavage 및 8cell 발달율 사이에는 유의적인 차이가 없었으나 배반포(BL) 및 8cell에서 배반포로 발달률은 18시간(BL 31$\pm$6, BL/8cell 82 $\pm$5%)에서 가장 높게 나타났으며 24시간(BL 17$\pm$2, BL/8cell 60$\pm$8%)과 유의적인 차이를 보였다(P<0.05). IVC 7일째 배반포의 총세포수와 trophoblast(TE) 세포수는 IVM 18시간(mean$\pm$S.E.; total: 131.1$\pm$34.0, TE: 97.6$\pm$29.6)에서 24시간(total: 112.2$\pm$17.5, TE: 80.1$\pm$15.6)보다 유의하게 많은 것으로 나왔으나(P<0.05) 7일째의 inner cell mass(ICM) 숫자(18hr 33.5$\pm$12.8 vs 24hr 32.1$\pm$12.0)와 8일째 ICM, TE 그리고 총 세포수에는 유의성 있는 차이가 없었다. IVM 18시간에서 PB 형성과 8cell 발달률 사이에 높은 상관성을 보였고 배반포 및 8cell에서 배반포 단계로 높은 발달률을 보였으며 생산된 배반포의 TE 숫자와 총 세포수가 유의하게 많은 것으로 나타났다. 따라서 IVM 18시간 실시하였을 경우 보다 많은 세포수를 가진 배반포 발달 가능성이 높은 embryo를 조기에 선발 가능할 것으로 사료된다.

• Clinical and Experimental Reproductive Medicine
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• v.40 no.2
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• pp.83-89
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• 2013

Objective: To investigate outcomes of stimulated IVF cycles in which GnRH antagonist was omitted on the ovulation triggering day. Methods: A total of 86 women who underwent controlled ovarian hyperstimulation with recombinant FSH and GnRH antagonist flexible multiple-dose protocols were recruited and prospectively randomized into the conventional group (group A) or cessation group (group B). The GnRH antagonist, 0.25 mg/day of cetrorelix, was started when the leading follicle reached 14 mm in diameter and was continuously administered until the hCG triggering day (group A, 43 cycles) or until the day before hCG administration (group B, 43 cycles). The maturity of oocytes, fertilization rate, embryo quality, and implantation and clinical pregnancy rates were evaluated. Results: The duration of ovarian stimulation, total dose of gonadotropins, serum estradiol levels on hCG administration day, and number of oocytes retrieved were not significantly different between the two groups. The total dose of GnRH antagonist was significantly lower in group B than group A ($2.5{\pm}0.9$ vs. $3.2{\pm}0.8$ ampoules, p<0.05). There was no premature luteinization in any of the subjects. The proportion of mature oocytes and fertilization rate were not significantly different in group B than group A (70.7% vs. 66.7%; 71.1% vs. 66.4%, respectively). There were no significant differences in the implantation or clinical pregnancy rates. Conclusion: Our prospective randomized study suggested that cessation of GnRH antagonist on the hCG administration day during a flexible multiple-dose protocol could reduce the total dose of GnRH antagonist without compromising its effects on pregnancy rates.

• Clinical and Experimental Reproductive Medicine
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• v.38 no.3
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• pp.148-152
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• 2011

Objective: Laser-assisted intracytoplasmic sperm injection (LA-ICSI), also known as micro-opening or thinning of the zona pellucida (ZP) prior to ICSI, may help to reduce mechanical damage to the oocyte during the procedure. The aim of the present study was to evaluate and analyze the efficacy of our institutional LA-ICSI program, which features laser-assisted ZP thinning prior to ICSI, in comparison with conventional ICSI (C-ICSI), performed on patients with different clinical characteristics. Methods: Patients undergoing a total of 212 ICSI cycles were randomly divided into an LA-ICSI group (106 cycles) and a conventional ICSI group (106 cycles). To reduce tissue damage, we thinned the ZP by approximately 70%, using a laser, before ICSI. Patients thus treated formed the LAICSI group. Comparisons included the morphological quality of transferred embryos, blastocyst development of the remaining embryos, and clinical pregnancy, in terms of ICSI method and patient characteristics. Results: Fertilization, development of remaining embryos, and pregnancy rate were significantly higher in the LA-ICSI group compared with the C-ICSI group. Fertilization, embryonic development, and the pregnancy rate were all improved in younger patients (<38 years of age) and in those who underwent a low number of IVF-ET attempts (<3 trials). In addition, the pregnancy rate was increased in older patients. Conclusion: LA-ICSI may be useful in improving the chance of pregnancy in all ICSI patients.

• Reproductive and Developmental Biology
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• v.30 no.3
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• pp.189-194
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• 2006

Sperm-mediated gene transfer(SMGT) can be used to transfer exogenous DNA into the oocyte at fertilization. The main objective of this study was to assess efficiency of transferring mitochondrial DNA(mtDNA) fragment into boar spermatozoa in either presence or absence of liposome and quality of transfected spermatozoa. The mtDNA of chicken liver was isolated and purified by phenol and alkaline lysis extraction, and it was inserted to plasmid. The genome of transfected spermatozoa treated with DNase I was purified by alkaline lysis, and then amplified by the PCR analysis. After electrophoresis, DNA quantitation of each well was calculated by comparison of the band intensity with standard. As a result, exogenous DNA was composed of mtDNA fragment(1.2 kb) and plasmid(2.7 kb). On the other hand, efficiency of transfection by liposome($9.0{\pm}0.34ng/{\mu}l$) in SMGT was higher than that by DNA solution($6.9{\pm}0.53ng/{\mu}l$). However, there was no significant difference. Transfering exogenous DNA into spermatozoa was completed within 90 min of incubation. In another experiment, there were significant (p<0.05) differences between transfected spermatozoa using both DNA solution and DNA/liposome completes with unheated spermatozoa for viability ($70.8{\pm}1.80$ and $68.0{\pm}2.16%$ vs. $83.3{\pm}1.69%$, respectively) and motility($78.7{\pm}1.59$ and $79.3{\pm}2.14%$ vs. $86.7{\pm}1.59%$, respectively). This study indicates that exogenous mtDNA can be efficiently transferred into boar spermatozoa regardless of the presence of liposome, and transfected spermatozoa can also use insemination and in vitro fertilization to generate transgenic pig.

• Journal of Animal Science and Technology
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• v.63 no.1
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• pp.1-15
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• 2021

As a result of intensive breeding, litter size has considerably increased in pig production over the last three decades. This has resulted in an increase in farrowing complications. Prolonged farrowing will shorten the window for suckling colostrum and reduce the chances for high-quality colostrum intake. Studies also agree that increasing litter sizes concomitantly resulted in decreased piglet birth weight and increased within-litter birth weight variations. Birth weight, however, is one of the critical factors affecting the prognosis of colostrum intake, and piglet growth, welfare, and survival. Litters of uneven birth weight distribution will suffer and lead to increased piglet mortality before weaning. The proper management is key to handle the situation. Feeding strategies before farrowing, management routines during parturition (e.g., drying and moving piglets to the udder and cross-fostering) and feeding an energy source to piglets after birth may be beneficial management tools with large litters. Insulin-like growth factor 1 (IGF-1)-driven recovery from energy losses during lactation appears critical for supporting follicle development, the viability of oocytes and embryos, and, eventually, litter uniformity. This paper explores certain management routines for neonatal piglets that can lead to the optimization of their colostrum intake and thereby their survival in large litters. In addition, this paper reviews the evidence concerning nutritional factors, particularly lactation feeding that may reduce the loss of sow body reserves, affecting the growth of the next oocyte generation. In conclusion, decreasing birth weight and compromised immunity are subjects warranting investigation in the search for novel management tools. Furthermore, to increase litter uniformity, more focus should be placed on nutritional factors that affect IGF-1-driven follicle development before ovulation.