• 한국수정란이식학회지
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• 제21권2호
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• pp.121-128
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• 2006

본 연구는 고품질의 돼지 체외 수정란을 생산하기 위하여 체외 성숙 배지에 첨가하는 polyvinylpyrrolidone(PVP)의 분자량, 첨가 농도 및 시간(실험 1)과 체외 성숙 수정 배양 단계에서 PVP의 첨가(실험 2)가 배 발생과 세포수에 미치는 효과를 검토하였다. 돼지 미성숙 난자의 체외 성숙은 NCSU 23 용액, 체외 수정은 mTBM 용액, 체외 배양은 PZM 3 용액을 이용하였다. 체외 성숙용 배지에서 PVP의 분자량, 농도 및 첨가 시간에 따른 수정율과 배반포 발달율은 차이가 없었다. 그러나 배반포의 부화율은 분자량 40,000(30.1%), 0.5%(28.6%) 및 $0{\sim}44$시간(37.5%) 첨가군이 다른 시험군에 비하여 유의하게 높았다(p<0.05). 분자량 40,000 0.5% PVP를 체외성숙, 수정 및 배양 배지에 각각 첨가(W) 또는 미첨가(W/O)한 결과, 수정율은 체외성숙 수정 배양에서 W-W/O-W 군이 69.3%, 배반포 발달율과 부화율은 W-W/O-W 및 W-W/O-W/O 군이 각각 12.6과 30.0% 및 14.0과 37.5%로서 다른 군에 비하여 유의하게 높았다(p<0.05). 총 세포 수와 Trophoblast 수는 처리군 간에 유사한 경향이었으나, Inner cell mass 수는 W/O-W/O-W 군이 평균 7.4개로서 가장 높았다 (p<0.05).

• 한국수정란이식학회지
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• 제13권1호
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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.

• 한국가축번식학회지
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• 제20권2호
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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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• 제42권2호
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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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• 제25권2호
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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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• 제18권2호
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• pp.197-203
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• 2011

목적: 35세 이상 산모에서 체외 수정 시술을 통해 출생한 쌍생아와 자연 임신 쌍생아의 임상 양상을 비교하여 35세 이상 산모와 체외 수정 시술로 출생한 신생아 관리에 기초 자료 제공을 위하여 실시하였다. 방법: 2001년 1월 1일부터 2010년 12월 31일 사이 분당차병원에서 출생한 쌍생아 중 산모의 나이가 35세 이상인 신생아 508례를 대상으로 하여, 체외 수정 시술을 시행 받은 288례와 자연 임신군 220례 사이의 주산기 특성과 합병증 및 신생아기 질환발생의 차이를 의무 기록을 통해 후향적으로 조사하였다. 결과: 체외 수정 시술군과 자연 수정군 사이의 산모 연령은 (36.7${\pm}$2.07세 vs. 36.8${\pm}$2.18세, P=0.57)로 통계적 차이는 보이지 않았다. 재태 연령($36^{+0}{\pm}1^{+5}$주 vs. $36^{+0}{\pm}2^{+0}$주, P=0.95), 출생체중(2,420${\pm}$440 g vs. 2,480${\pm}$460 g, P=0.14) 역시 차이는 보이지 않았다. 1분 아프가 점수(7.37${\pm}$1.19 vs. 7.09${\pm}$1.46, P=0.019)와 5분 아프가 점수(8.67${\pm}$0.84 vs. 8.51${\pm}$0.96, P=0.045)는 모두 체외 수정 시술군에서 높았다. 임신성 당뇨, 임신성 고혈압, 전치태반, 조기 양막 파수, 제왕 절개술, 부당 경량아의 빈도는 두 군간의 차이가 없었다. 조발형 패혈증의 빈도는 체외 수정 시술군에서 자연 수정군보다 낮았다(2.4% vs. 6.4%, P=0.02). 그밖에 신생아 호흡 곤란 증후군, 기관지 폐 형성이상, 동맥관 개존증, 신생아 괴사 장염, 뇌실 내 출혈의 발생 빈도는 모두 두 군 간에 차이가 없었다. 결론: 본 기관에서 지난 10년간 고연령 산모에서 체외 수정 시술로 태어난 쌍생아의 임상 양상을 조사하여 자연 수정으로 출생한 쌍생아와 비교하였을 때, 체외 수정 시술로 인하여 주산기 합병증과 신생아기 질환의 발생 빈도가 높아지지는 않았다.

• 한국수정란이식학회지
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• 제18권3호
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• pp.237-242
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• 2003

본 연구에서는 소에 있어서 수정란 이식을 다각적으로 분석하고 개선하기 위해 이식한 결과를 비교 검토하였고 그 결과는 다음과 같다. 1. 채란된 한우 난포란을 이용하여 생산된 체외수정란을 서로 다른 지역별로 이식한 바 B지역(경주)에서는 106의 수란우에 이식하여 51두(48.1％)가 수태하였고, A지역(김천) 수태율(33.8％)과 D 지역(경산) 수태율(35.3％)보다 유의하게 높았지만, C 지역(탑리)에서는 유의차가 존재하지 않았다. 따라서, 수정란이식에 있어서 이식 지역과 사육 환경, 시술자의 숙련도에 따라 수태율이 달라질 수 있다는 것을 알 수 있었다. 2. 수란우의 산차에 따른 수태율은, 미경산우 42.9％로서 경산우의 36.6％에 비해 다소 높은 수태율을 보였으나 유의적인 차이는 인정되지 않았다(P<0.05). 3. 이식 수정란의 발육단계는 중기 배반포 또는 후기 배반포로서 구분하여 수란우에 이식하였다. 중기 배반포가 45.5％, 후기 배반포 41.0％로서 중기 배반포가 후기 배반포보다 높은 수태율을 나타내었지만 유의한 차이는 인정되지 않았다. 배반포기가 생성된 날짜에 따른 수란우의 수태율은 7일째 생성된 배반포 이식시 수태율은 43.8％, 8일째 생성된 배반포 이식시 수태율은 32.9％, 9일째 생성된 배반포 이식시 수태율은 20.0％로서 7일째 배반포의 수태율 이 다른 두 처리군보다 수태율은 높았지만, 유의차는 인정되지 않았다.

• 한국가축번식학회지
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• 제24권3호
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• pp.311-317
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• 2000

본 연구는 체외에서 한우 난포란의 성숙과 수정후 배의 체외발생에 있어서 온도충격의 영향이 처리온도 및 처리시간에 미치는 효과를 검토하였다. 1. 체외에서 생산된 4~8 세포기 배의 배반포로의 배발달율에 있어서 온도충격의 적정온도는 41$^{\circ}C$, 처리시간은 30초였다. 2. 체외성숙 후의 온도충격이 체외성숙 전의 온도충격보다 수정란의 분할율을 유의하게 상승시켰다(p<0.05). 3. 체외성숙 후와 배양 5일째 배에 각각 온도처리를 하였을 경우 비처리군보다 배반포의 배발달율이 향상되었다.

• Clinical and Experimental Reproductive Medicine
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• 제48권4호
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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.

• 한국수정란이식학회지
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• 제32권3호
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• pp.123-130
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• 2017

This study was conducted to investigate the effects of alpha-lipoic acid (aLA) as an antioxidant that decrease the reactive oxygen species (ROS) in bovine embryonic development. Slaughterhouse derived bovine immature oocytes were collected and 4 different concentrations (0, 5, 10 and 20 mM) of aLA was supplemented in bovine in vitro maturation (IVM) medium. After 20 hrs of IVM, maturation rates, levels of ROS and glutathione (GSH), and further embryonic development after parthenogenetic activation (PA) and in vitro fertilization (IVF) was investigated according to aLA concentrations. Maturation rate was significantly higher in 10 mM group than other groups (80.5% vs. 62.9, 73.9, 64.2%; P<0.05). In the levels of ROS and GSH in matured oocytes as an indicator of oocyte quality, significantly better results were shown in 5 and 10 mM groups compared with other 2 groups. After IVM, significantly higher rates of blastocyst formation were shown in 10 mM groups in both of PA (27.9% vs. 18.8, 22.3, 14.2%; P<0.05) and IVF (32.6% vs. 23.9, 27.3, 16.2%; P<0.05) embryos. In addition, significantly more cell total cell number and higher inner cell mass ratio in 10 mM PA and IVP blastocysts showed developmental competence in 10 uM groups. Therefore, based on the entire data from this study, using $10{\mu}M$ of aLA confirmed to be the optimal concentration for bovine oocyte maturation and embryonic development.