• Asian-Australasian Journal of Animal Sciences
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• 제20권10호
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• pp.1510-1516
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• 2007

These experiments were carried out to optimize the parameters of electrical activation, methods of parthenogenetic activation and embryo culture in vitro and meanwhile to isolate embryonic stem cells-like (ESCs) derived from porcine parthenogenetic blastocysts (pPBs). These results showed that, as the electric field strength increased from 1.0 to 2.7 kV/cm, the cleavage rate of parthenogenetic embryos increased gradually but the rate of oocyte lysis was significantly increased when using 2.7 kV/cm field strength. The rate of cleavage in 2.2 and 2.7 kV/cm groups was significantly increased in comparison with that of the 1.0 kV/cm group. A voltage field strength of 2.2 kV/cm DC was used to investigate blastocyst development following activation with a single pulse of 30 or $60-{\mu}sec$ pulse duration. The optimum pulse duration was 30-${\mu}sec$, with a blastocyst rate of 20.7%. Multiple pulses were inferior to a single pulse for blastocyst yield (8.0% vs. 29.9) (p<0.05). For porcine oocyte parthenogenetic activation methods, the rates of cleavage (79.0% vs. 59.8%) and blastocysts (19.4% vs. 3.4%) were significantly increased in electrical activation in contrast to chemical activation with ionomycin/6-DMAP (p<0.05). Rates of cleavage and blastocyst formation in NCSU-23 and PZM-3 embryo media were higher than those of G1.3/G2.3 serial culture media, but there was no significant difference among the three groups. The total cell number of blastocysts in PZM-3 embryo culture media containing $5{\mu}g/ml$ insulin was significantly higher than that of the control (no insulin) ($44.3{\pm}9.1$ vs. $33.9{\pm}11.7$). For isolation of PESCs-like, the rates of porcine blastocysts attached to feeder layers and ICM colony formation in Method B (nude embryo culture) were better than those in Method A (intact embryo culture).

• Clinical and Experimental Reproductive Medicine
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• 제28권4호
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• pp.307-315
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• 2001

Objective: To evaluate the clinical outcomes and influencing factors of in vitro fertilization and embryo transfer (IVF-ET) in patients with failed pregnancy after microsurgical reversal of tubal sterilization. Materials and Methods : From January, 1997 to December, 2000, IVF-ET was performed in two groups; the study TR (tubal reanastomosis) group consisted of 147 cycles in 66 patients with failed microsurgical reversal of tubal sterilization, and the control group of 115 cycles in 67 patients with bilateral tubal occlusion (BTO). The two groups were evaluated and compared for clinical characteristics, clinical pregnancy rates, and factors influencing the outcomes of IVF-ET. Results: Compared with the control BTO group, age and the previous parity were significantly higher ($36.3{\pm}2.7$ vs. $33.6{\pm}2.0$ years, p<0.05; $1.6{\pm}0.7$ vs. $0.2{\pm}0.4$, p<0.05), and the clinical pregnancy rate per cycle was significantly lower (23.8% (35/147) vs. 29.3% (34/115), p<0.05) in the TR group. Difference in the clinical pregnancy rates was age-related, since there was no significant difference between the two groups, except for the previous parity ($1.6{\pm}0.7$ vs. $0.1{\pm}0.3$, p<0.05), when the patients aged 37 years or older were excluded. No difference was found in terms of the following: the proportion of controlled ovarian hyperstimulation (COH) cycles with GnRH agonist ultrashort protocol, the duration of COH, the dosage of gonadotropins used, and the numbers of oocytes retrieved and of embryos transferred, irrespective of age correction. Conclusions: The outcomes of IVF-ET following the failed microsurgical reversal of tubal sterilization depend upon patient age. The previous fertility of patients does not seem to be a factor of better IVF-ET prognosis.

• Asian-Australasian Journal of Animal Sciences
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• 제13권5호
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• pp.587-594
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• 2000

At present embryonic stem (ES) cells with confirmed pluripotential properties are only available in the mouse. Recently, we were able to isolate, culture and genetically transform primordial germ cell (PGC)-derived cells from pig embryos and demonstrate their ability to contribute to chimera development in the pig. In order to determine whether the system we developed could be used to isolate embryonic germ (EG) cells from other mammalian species, we placed isolated PGCs from cattle, goats, rabbits and rats in culture. Briefly, PGCs were isolated from fetuses of cow (day 30-50), goat (day 25), rabbit (day 15-18) and rat (day 11-12), and plated on STO feeder cells in Dulbecco's modified Eagle's medium (DMEM): Ham's F10 medium (1:1) supplemented with 0.01 mM nonessential amino acids, 2 mM L-glutamine, 0.1 mM $\beta$ - mercaptoethnol, soluble recombinant human stem cell factor (SCF; 40ng/ml), human basic fibroblast growth factor (bFGF; 20ng/ml) and human leukemia inhibitory factor (LIF; 20ng/ml). For maintenance of the cells, colonies were passed to fresh feeders every 7-10 days. In all species tested, we were able to obtain and maintain colonies with ES-like morphology. Their developmental potential was tested by alkaline phosphatase (AP) staining and in vitro differentiation assay. For genetic transformation, cells were electroporated with a construct containing the green fluorescent protein (GFP) under the control of the cytomegalovirus (CMV) promoter. GFP-expressing colonies were detected in cattle, rabbits and rats. These results suggest that PGC-derived cells from cattle, goats, rabbits and rats can be isolated, cultured, and genetically transformed, and provide the basis for analyzing their developmental potential and their possible use for the precise genetic modification of these species.

• 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.

• Clinical and Experimental Reproductive Medicine
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• 제37권1호
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• pp.33-39
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• 2010

목 적: 체외수정 및 배아이식술에서의 출생 성비는 사회적으로 과학적으로도 매우 관심이 있는 분야이지만 국내에서는 아직까지 정확하게 조사된 바가 없다. 본 연구에서는 국가 통계에서 출생 성비의 추이를 살펴보고, 국내 체외수정 및 배아이식술에서의 출생 성비에 대해 조사하고자 하였다. 연구방법: 출생 성비 국가 통계 (1991~2008)는 통계청 자료를 통해 확인하였으며, 체외수정의 분만 자료 (2007~2008)는 보건복지가족부의 용역 연구 결과를 제공받아 분석하였다. 체외수정 방법에 따라 일반적인 체외수정과 세포질내 정자주입술 그리고 이식된 배아에 따라 신선 주기 배아이식과 동결-해동 배아이식으로 구분하여 출생 성비를 통계학적인 방법으로 비교하였다. 결 과: 우리나라의 출생 성비는 2002년까지 불균형 상태인 1.10 정도로 유지되었으나, 그 이후 감소되기 시작하여 2007년과 2008년에는 균형 출생 성비인 1.06이 유지되고 있으며, 체외수정에서의 출생 성비도 2007년 1.07, 2008년 1.06을 나타내었다. 체외수정 방법에 따른 출생 성비는 2008년의 일반적인 체외수정에서 1.10로 가장 높았으며, 2008년의 세포질내 정자주입술에서 1.01로 가장 낮게 나타났다. 체외수정 방법과 이식배아에 따른 각 군간의 비교에서 통계적인 유의성은 나타나지 않았다. 결 론: 국내에서 처음으로 조사된 2007년과 2008년 체외수정 및 배아이식술의 출생 성비도 각각 1.07과 1.06으로 우리나라의 전체적인 출생 성비와 매우 유사하였으며, 정상적인 출생 성비를 나타냈다. 향후 국내의 전체적인 체외수정 및 배아이식술에 대한 통계와 다양한 요인들에 따른 보다 전향적인 출생 성비에 대한 연구가 필요할 것으로 생각된다.

• Journal of Nutrition and Health
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• 제57권1호
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• pp.53-64
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• 2024

본 연구는 근생성 효능을 가진 천연화합물을 발굴하고자 oxypeucedanin hydrate가 근생성과 미토콘드리아에 미치는 영향 및 항암제 유도 미토콘드리아 손상에 대한 완화효과를 C2C12 근원세포와 zebrafish 모델을 통해 각각 확인하였다. 그 결과, oxypeucedanin hydrate는 다핵의 근관세포의 수와 분화말기 표지자인 Myh4의 발현량을 증가시켰고 근육단백질 분해 인자인 MuRF1과 MAFbx의 발현량은 감소시켰다. 또한 미토콘드리아 생합성 조절인자인 Pgc1α, Tfam과 전자전달계 구성인자인 Sdha, Cox1의 발현은 증가시키고, 미토콘드리아 융합인자인 Opa1의 발현 또한 증가시킴과 동시에 미토콘드리아 분열을 표지하는 Drp1의 발현은 감소시켰다. 한편 zebrafish 모델을 통해 항암제 유도 미토콘드리아 손상에 대한 개선효과를 확인한 결과, oxypeucedanin hydrate는 항암제에 의해 유도된 미토콘드리아 손상을 완화시켰다. 이상의 결과들은 oxypeucedanin hydrate가 미토콘드리아 기능 증진을 매개로 근원세포 분화 촉진 및 근육 단백질 분해 저하 효과를 나타냄을 시사한다. 따라서 본 연구를 통해 oxypeucedanin hydrate가 근생성 효과를 나타낼 수 있는 잠재적인 유효소재로서의 가능성을 제시하였다.

• Asian-Australasian Journal of Animal Sciences
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• 제28권12호
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• pp.1703-1712
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• 2015

The in vitro maturation (IVM) efficiency of porcine embryos is still low because of poor oocyte quality. Although brilliant cresyl blue positive (BCB+) oocytes with low glucose-6-phosphate dehydrogenase (G6PDH) activity have shown superior quality than BCB negative (-) oocytes with high G6PDH activity, the use of a BCB staining test before IVM is still controversial. This study aimed to shed more light on the subcellular characteristics of porcine oocytes after selection using BCB staining. We assessed germinal vesicle chromatin configuration, cortical granule (CG) migration, mitochondrial distribution, the levels of acetylated lysine 9 of histone H3 (AcH3K9) and nuclear apoptosis features to investigate the correlation between G6PDH activity and these developmentally related features. A pattern of chromatin surrounding the nucleoli was seen in 53.0% of BCB+ oocytes and 77.6% of BCB+ oocytes showed peripherally distributed CGs. After IVM, 48.7% of BCB+ oocytes had a diffused mitochondrial distribution pattern. However, there were no significant differences in the levels of AcH3K9 in the nuclei of blastocysts derived from BCB+ and BCB- oocytes; at the same time, we observed a similar incidence of apoptosis in the BCB+ and control groups. Although this study indicated that G6PDH activity in porcine oocytes was correlated with several subcellular characteristics such as germinal vesicle chromatin configuration, CG migration and mitochondrial distribution, other features such as AcH3K9 level and nuclear apoptotic features were not associated with G6PDH activity and did not validate the BCB staining test. In using this test for selecting porcine oocytes, subcellular characteristics such as the AcH3K9 level and apoptotic nuclear features should also be considered. Adding histone deacetylase inhibitors or apoptosis inhibitors into the culture medium used might improve the efficiency of IVM of BCB+ oocytes.

• 한국가축번식학회지
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• 제22권4호
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• pp.395-403
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• 1998

본 연구에서는 돼지난자내 원형정자 주입후 수정율과 체외 배 발달을 조사하였다. 원형정자 주입 2시간 전에 인위적 전기자극을 주고 원형정자를 주입했을 때 난자들의 수정율이 주입 직후에 전기자극을 준 것들과 전기 자극을 주지 않은 것들보다 수정율이 높았다. 원형정자와 원형 정자핵을 각각 주입 한 후 전핵 형성율과 전핵 이동율을 조사하였으나 유의차를 발견할 수 없었다. 전핵의 형성과 이동중 미세소관의 움직임을 간접형광면역법 및 공춧점 현미경을 사용하여 조사해본 결과, 난활성 직후 난자의 표층에서 미세소관이 발생해서 이것에 의해 웅성 및 자성전핵이 난자 중심부로 이전됨을 볼 수 있었다. 원형정자와 원형정자핵 주입후 배양 6일째에 각각 25% 와 27%의 배반포로 형성되었고, 8 일째 형성된 배반포를 염색하여 관찰한 철과 세포수가 각각 평균 87에서 99개가 형성되었음을 알 수 있었다. 이러한 결과는 체외 성숙된 돼지 난자 내에 원형정자 혹은 원형정자 핵을 미세 주입하는 방법에 의해 정상척인 배발달을 하는 돼지 수정란 생산이 가능한 것을 보여주는 것이다.

• 한국가금학회:학술대회논문집
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• 한국가금학회 2005년도 제22차 정기총회 및 학술발표회
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• pp.66-67
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• 2005

배자 생식세포 발달에 관련된 메카니즘을 밝혀내기 위해서, 닭 배자 생식기에서 추출한 원시 생식세포의 단백질체 지도를 만들었다. 총 500 배자를 6일간 배양하여 배자 생식기를 획득했고, 7-10일 배양 후, 배양된 원시생식세포는 2차원 젤 전기 영동법에 의해 분할되어 졌다. 유의적 발현 수준을 나타낸 많은 단백질 스팟 들은 MALDI-TOP 와 LC-MS/MS에 의해 확인되었으며, 89개의 단백질 스팟 중에 50개의 mass spectra 들이 데이터베이스에서 조류 단백질과 일치함을 확인하였다. 본 실험에서 행한 단백질체 지도는 형질전환 연구와 생식세포 생물학 분야에 중요한 참고 문헌으로 가치를 가질수 있을 것이다.

• 한국약용작물학회지
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• 제11권5호
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• pp.336-339
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• 2003

약용식물로 쓰이는 선피막이 (Hydrocotyle maritima Honda)를 기내에서 재분화 가능성에 대해 조사하였다. 선피막이의 엽병 절편체를 식물생장조절제 ($0{\sim}5\;mg/l$, NAA와 $0{\sim}5\;mg/l$ 2,4-D)가 단독 또는 $0.1{\sim}2\;mg/l$ BA와 조합 첨가된 배지 에서 6주동안 배양하였다. 2,4-D 또는 NAA을 단독으로 처리해도 캘러스가 발생하나, 가장 좋은 결과는 각각 $0{\sim}5\;mg/l$ BA, $5\;mg/l$ NAA와 $0.5\;mg/l$ BA조합구에서 나타났다. Kinetin를 $3\;mg/l$로 처리시 캘러스로부터 가장 많은 신초 (캘러스당 12개)을 발생시켰다. 또한, $0.5\;mg/l$ 2,4-D와 $0.5\;mg/l$ BA가 첨가된 배지에서 유도된 배 발생 캘러스를 호르몬이 없는 배지에서 배양하였을 때, 체세포 배가 분화되었고, 식물체로 더 잘 발달하였다.