• Clinical and Experimental Reproductive Medicine
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• 제42권3호
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• pp.118-125
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• 2015

Objective: The goal of this study was to evaluate the pregnancy potential of immature (metaphase I or germinal vesicle stage) oocytes retrieved in intracytoplasmic sperm injection (ICSI) cycles. Methods: A total of 1,871 couples with infertility underwent 2,984 ICSI cycles. Cycles in which three or fewer oocytes were retrieved were included in this study in order to evaluate the pregnancy potential of immature oocytes. Cycles were divided into five groups (group I-V), according to the maturation status of the oocytes at the time of cumulus cell removal and ICSI. The fertilization and pregnancy rates after ICSI were analyzed and compared among the study groups based on the maturation status of the retrieved oocytes. Results: The retrieval of only immature oocytes was associated with a significant decrease in the fertilization rate ($76.1%{\pm}37.3%$ vs. $49.0%{\pm}49.1%$, $66.7%{\pm}48.7%$; group I vs. group II, group III, respectively) and the average number of transferred embryos ($1.5{\pm}0.7$ vs. $1.1{\pm}0.4$, $1.1{\pm}0.6$). The cycle cancellation rate was significantly higher when only immature oocytes were retrieved. The clinical pregnancy rate decreased significantly when the transferred embryos had originated from immature oocytes (16.9% vs. 10.3%, 1.2%). Conclusion: In ICSI cycles, the fertilization potential and pregnancy potential of the immature oocytes retrieved in ICSI cycles were inferior to those of mature oocytes. Therefore, increasing the number of injectable oocytes and transferrable embryos by using immature oocytes after their spontaneous in vitro maturation does not necessarily improve pregnancy outcomes.

• Clinical and Experimental Reproductive Medicine
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• 제31권3호
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• pp.155-168
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• 2004

Objective: Melatonin, which is secreted by pineal gland play an important role in the regulation of ovarian function via seasonal rhythm and sleep in most mammals. It also has a role in the protection of cells by removing toxic oxygen free radicals brought about by metabolism. In the present study, effects of melatonin on the mouse oocyte maturation were examined using two different culture conditions provided with 5% or 21% oxygen concentration. Material and Method: Immature mouse oocytes were obtained from the ovarian follicles of $3{\sim}4$ weeks old ICR strain mice intraperitoneally injected with 5 I.U. PMSG 44 hour before. Under stereomicroscope, morphologically healthy oocytes with distinct germinal vesicle (GV) were liberated from the graafian follicles and collected using mouth-controlled micropipette. They were then cultured for 17 hour at $37^{circ}C$, 5% $CO_2$ and 21% $O_2$ (95% air) or 5% $CO_2$, 5% $O_2$ and 90% $N_2$. New modified Hank's balanced salt solution (New MHBS) was used as a culture medium throughout the experiments. Effects of melatonin were examined at a concentration of $0.0001{\mu}M$, $0.01{\mu}M$ or $1.0{\mu}M$. For the prevention of spontaneous maturation of immature oocytes during culture, dibutyryl cyclic AMP (dbcAMP) and/or hypoxanthine were included in the medium. Results: Under 21% oxygen condition, oocytes cultured in the presence of $0.01{\mu}M$ melatonin showed a significantly higher maturation rates, in terms of germinal vesicle breakdown (95.0% vs 89.0%) and polar body formation (88.1% vs 75.4%), compared to those cultured with $0.0001{\mu}M$ or $1.0{\mu}M$ melatonin. However, no difference was observed in oocytes cultured under 5% oxygen whether they were treated with melatonin or not. In the presence of $0.01{\mu}M$ melatonin, oocytes either cultured under 21% or 5% oxygen exhibited no difference in the polar body formation (85.6% vs 86.7%). However, in the absence of melatonin, oocytes cultured under 21% oxygen exhibited lower polar body formation (74.7%). When oocytes were cultured in the presence of dbcAMP alone or with varying concentrations of melatonin, those treated with both compounds always showed better maturation, i.e., germinal vesicle breakdown and polar body formation, compared to those cultured with dbcAMP alone. At the same concentration of melatonin, however, oocytes exposed to 21% oxygen showed poor maturation than those to 5% oxygen. Similar results were obtained from the experiments using hypoxanthine instead of dbcAMP. Conclusion: Based upon these results, it is suggested that melatonin could enhance the meiotic maturation of mouse oocytes under 21% oxygen concentration, and release oocytes from the meiotic arrest by dbcAMP or hypoxanthine regardless of the concentration of oxygen, probably via the removal of oxygen free radicals.

• 한국수정란이식학회지
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• 제18권1호
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• pp.69-73
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• 2003

본 연구에서는 TCM-199, mSOF, NCSU-23 세 종류의 배지의 자연 단위 발생 유도로 형성된 돼지 배아의 초기 발육에 대한 효과를 조사하였다. 실험 1에서는 세 종류의 배지에서 체외 성숙된 도축장 유래 돼지 난자의 성숙율을 성숙 48 시간째 핵상 관찰로 조사하였다. 각 배지에서의 핵 성숙율은 mSOF군에서 83.1$\pm$2%, NCSU-23 군에서 78.0$\pm$3%, TCM-199군에서 83.5$\pm$2%로 세 배지의 난자 성숙율에 대한 유의적 차이는 없었다(P(0.05). 실험 2에서는 돼지 난자의 전핵 형성률, 단위 발생으로 형성된 배아의 분할율, 6-8세포기까지의 발육율 등이 각각 체외 성숙 55-58 시간, 96시간 그리고 168 시간 후에 관찰되었다. 전핵 형성율 (5.4$\pm$2 % in mSOF and 3.7$\pm$1% in NCSU-23 vs 1.7$\pm$3% in TCM-199) 그리고 6-8cell까지의 발육율은 (3.2$\pm$3% in mSOF and 4.0$\pm$1% in NCSU-23 vs 1.4$\pm$3% in TCM-199) mSOF군과 NCSU-23군에서 유의적으로 높았다(P<0.05). 이상의 결과를 볼 때 mSOF 배지와 NCSU-23 배지에서 장시간 배양에 의한 난자의 단위발생 유도율이 유의적으로 높았다.

• 한국발생생물학회지:발생과생식
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• 제17권2호
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• pp.141-147
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• 2013

Keeping the intact germinal vesicle (GV) is essential for maintaining the capacity of mammals including human. It is maintained by very complex procedures along with folliculogenesis and is a critical step for getting competent oocyte. So far, a few mechanisms involved in folliculogenesis are known but GV arrest mechanisms are largely unrevealed. Cyclic AMP, a adenosine derived substance, have been used as inhibitor of germinal vesicle breakdown as a putative oocyte maturation inhibitor. In this study, we examined the potency of adenosine as GV maintainer and a possible signaling mediator for that. A1, A2b, and A3 were detected in cumulus cells of cumulus enclosed-oocyte (CEO). Intact of germinal vesicle was not kept like in follicle but the spontaneous maturation was inhibited by exogenous adenosine. It is inhibited with concentration dependent manners. Intracellular calcium level of cumulus was extensively increased after adenosine treatment. Based on these results it is suggested that one of the pathway for GV arrest by adenosine and its receptors is calcium mediated signaling pathway in CEO.

• Asian-Australasian Journal of Animal Sciences
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• 제17권3호
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• pp.317-324
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• 2004

The present study was conducted to investigate the involvement of nitric oxide (NO) in cumulus expansion, oocyte mortality and meiotic maturation of porcine cumulus enclosed oocytes (CEOs) cultured in two different models when gonadotropins, including follicle-stimulating hormone (FSH) and human chorionic gonadotropin (hCG) were presented or not. And the interaction between NO and $\beta$-mercaptoethanol ($\beta$-ME), a free radical scavenger was also investigated. Two models refer to spontaneous maturation model and hypoxanthine (HX) medium model. All the 3,433 eligible CEOs were incubated at $39^{\circ}C$ and the cumulus expansion, oocyte morphology and nuclear phase were evaluated 44 h after incubation. (1) In spontaneous maturation model, NO stimulates the cumulus expansion and $\beta$-ME delayed it. NO doesn't affect the oocyte meiotic resumption but inhibits the oocytes to develop to metaphase II. (2) In HX medium model, NO or $\beta$-ME doesn't affect the expansion in the absence of gonadotropins, but in the presence of gonadotropins, NO or $\beta$-ME inhibits the expansion. In the presence of gonadotropins, NO inhibits the oocyte meiotic resumption and it especially inhibits the oocyte to develop to metaphase II, and $\beta$-ME reverses such inhibitory effects. The cooperation of gonadotropins and $\beta$-ME stimulates the meiotic resumption and especially, promotes the CEOs to develop to metaphase II in both models. Moreover, HX might contribute to the fragility of oocyte zona pellucida and gonadotropins, nitric oxide and $\beta$-ME could alleviate it separately, and cooperatively. It is concluded that NO exerts different functions in two models and $\beta$-ME affected the functions of NO in different models.

• Reproductive and Developmental Biology
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• 제35권4호
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• pp.499-502
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• 2011

Aurora A kinase is a mitotic serine/threonine kinase whose proposed functions include the maturation of centrosomes, G2/M transition, alignment of chromosomes at metaphase, and cytokinesis. In this study, we investigated the effect of MLN8237, an aurora A kinase inhibitor, on the postovulatory aging of oocytes based on the frequency of oocyte fragmentation, cdk1 kinase activity, and cyclin B degradation. The fragmentation of ovulated oocytes during prolonged culture was inhibited by treatment with MLN8237 in a concentration-dependent manner. The frequency of fragmented oocytes was significantly lower in oocytes treated with 2 ${\mu}M$ MLN8237 (13%) than in control oocytes (64%) after two days of culture. Most of the control (non-fragmented) oocytes (91%) were activated after two days of culture. In comparison, only 22% of the MLN8237-treated oocytes were activated; the rest of the oocytes (78%) were still in metaphase with an abnormal spindle and dispersed chromosomes. Next, cdk1 activity and the level of cyclin B were examined. The level of cyclin B and cdk1 activity in MLN8237-treated oocytes were nearly equal to those in control oocytes. Our results indicate that MLN8237 inhibited the fragmentation of ovulated oocytes during prolonged culture, although it blocked the spontaneous decrease in activity of cdk1 and degradation of cyclin B. This mechanism of inhibition is different from that in oocytes treated with nocodazole, which have high levels of cdk1 activity and cyclin B.

• 한국동물번식학회:학술대회논문집
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• 한국동물번식학회 2003년도 학술발표대회 발표논문초록집
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• pp.61-61
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• 2003

We have investigated the novel function of tissue transglutaminase (tTG) in the germinal vesicle breakdown (GVBD) of mouse oocyte. tTG was identified in ooplasm and germinal vesicle by immunostaining assay. Spontaneous maturation of the oocytes elevated in situ activity of tTG by over 2.5 fold at 3 hr, which was determined by a confocal microscopic assay. However, incubation with monodansylcadaverine (MDC), a tTG inhibitor, blocked the activation of tTG. The possible role of tTG in GVBD was investigated by the use of two tTG inhibitors, MDC and cystamine. MDC largely inhibited the GVBD by a concentration dependent manner. GV-stage oocytes were matured to the GVBD stage by 78% at 3 hr in BWW culture medium. However, in the oocytes incubated with MDC for 3 hr, the GVBD rates were 43 and 11% by 50 and 100 mM, respectively. MDC also blocked the entry of 70 kDa TRITC-dextran from the ooplasm to the compartment of germinal vesicle, indicating a possible inhibition of nuclear pore disassembly by MDC. The role of tTG in GVBD was further investigated by microinjection with cystamine. The control oocytes, injected with DPBS, showed about 80 % of GVBD at 3 hr. But the oocytes injected with cystamine showed 15% of GVBD at 3 hr and a little higher rate at 6 hr. In addition, the inhibition of GVBD maturation by MDC was reversible by washing. These results suggested that tTG was involved in the early event of mouse oocyte maturation

• 한국동물학회지
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• 제31권2호
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• pp.87-94
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• 1988

전라남도 일대에서 서식하는 북방상개구리(R. dybowskii)와 참개구리(R. nigromaculara)를 채집하는 생체외배양에 의한 여포난자의 성숙을 유도하였다. 북방산개구리의 여포난자는 배양액(amphibian Ringer's soluion AR)에 첨가한 progesterone, 0.1 $\mu$g/2 ml에 의해 난자의 성숙(핵붕괴)이 유도 되었으며 참개구리의 난자는 1 $\mu$g/2 ml (frog pituitary homogenate FPH)을 얻어서 그 효과를 조사해본 결과 북방산개구리에서는 0.01 pituitary equivalent/2 ml에서, 참개구리는 0.1 pit equiv./2 ml에서부터 여포난자의 성숙이 일어났다. 난자의 성숙에 요하는 시간은 두 개구리에서 모두 9-15시간이었으며 호르몬에 대한 반응성, 성숙기간 등은 개구리 재료로 가장 많이 사용되는 법개구리(R. pipiens)와 거의 일치하였다. 특이하게 2개월 이후에 사용한 북방산개구리의 여포난자는 호르몬의 도움없이도 성숙이 일어났으며 성숙기간도 3시간으로 매우 빨라졌다. 난소조각을 배양했을 때 자발적으로 성숙을 일으키는 여포들은 자발적인 배란까지도 일으키는 것을 발견하였다.

• 한국양식학회지
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• 제14권4호
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• pp.213-220
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• 2001

본 연구에서는 농어과, 농어속 어류인 농어(L. japonicus)어미를 대상으로 2회에 걸쳐 성 성숙 및 자연 산란유도에 있어 염분의 영향을 조사하여 인위적 성숙 및 산란 조절가능성을 파악하였다. 실험기간 동안의 평균수온은 1차 실험의 대조구에서 14.3$\pm$1.1$^{\circ}C$, 저염분구에서 14.2$\pm$1.1$^{\circ}C$였고, 2차 실험의 대조구에서 15.8$\pm$1.2$^{\circ}C$, 저염분구에서 15.4$\pm$1.1$^{\circ}C$로 나타나 실험구에셔 15.8$\pm$1.2$^{\circ}C$, 저염분구에서 15.4$\pm$1.1$^{\circ}C$로 나타나 실험구간 유의차는 없었다. 그리고 성 성숙과 산란시기로 판단되는 11~2월 사이의 월별 평균수온은 1차 실험의 대조구에서 각각 10.6$\pm$0.8$^{\circ}C$~13.8$\pm$0.4$^{\circ}C$였고, 저염분구에서 각각 10.6$\pm$0.8$^{\circ}C$~14.6$\pm$1.4$^{\circ}C$로 나타났다. 2차 실험의 실험구 I은 각각 10.7$\pm$0.5$^{\circ}C$~15.8$\pm$1.$0^{\circ}C$였고, 실험구 II는 각각 10.3$\pm$0.9$^{\circ}C$~15.7$\pm$0.7$^{\circ}C$로 나타났다. 실험기간 동안의 평균염분은 1차 실험의 대조구에서 35.1$\pm$0.2$\textperthousand$, 저염분구에서 33.8$\pm$0.6$\textperthousand$였고, 2차 실험의 대조구에서 34.5$\pm$0.4$\textperthousand$, 저염분구에서 월별 평균염분은 1차 실험에서 각각 31.0$\pm$3.7~35.2$\pm$0.7$\textperthousand$(최저염분은 26.5$\textperthousand$)였고, 2차 실험에서 각각 24.8$\pm$0.9~35.1$\pm$0.7$\textperthousand$(최저염분은 23.4$\textperthousand$)였다. 각 실험구별 개체당 일간 사료섭식량은 1차와 2차 실험에서 저염분구가 대조구보다 많은 사료를 섭식했고, 또한 성장률 및 생존율도 두 번의 실험 모두에서 저염분구가 해수만을 공급한 대조구에 비해 높았다. 어미의 성 성숙 및 산란은 두 번의 실험에서 대조구보다 저염분구에서 원만히 이루어졌다. 암컷 성숙 개체의 경우 1차 실험은 대조구 6마리, 저염분구 12마리였으며, 2차 실험은 대조구 5마리, 저염분구 12마리였으며, 2차 실험은 대조구 5마리, 저염분구 14마리로서 성숙유도에 있어 염분의 조절에 의한 성숙이 이루어진 것을 알 수 있다. 산란 시기는 1차 실험에서 대조구나 저염분구의 산란 개시 시점이 거의 동일한 데 비해, 2차 실험에서는 저염분구가 대조구에 비해 대략 20일 정도 빠르게 나타났다. 또한 산란에 가입한 암컷 어미의 개체수도 두 차례의 실험 모두 저염분구가 많았다. 또한 알의 크기 면에서도 저염분구가 대조구의 53.8$\pm$4.6%을 나타낸 반면 대조구는 19.3$\pm$2.3%로 낮았고, 2차 실험에서도 대조구에서 산란된 알의 대부분이 과숙상태여서 정상적인 부화가 이루어지지 못한데 비해 저염분구의 알들은 56.4$\pm$7.1%의 부화율을 나타내었다.

• 한국공간정보시스템학회 논문지
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• 제3권1호
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• pp.109-116
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• 2001

ITS산업은 정보경제의 일반적 특징이라고 할 수 있는 네트워크 외부성과 정보의 양방향성을 갖고있는 동시에, 정보의 적시성과 지역성, 자연 독점성, 공공성, 시스템 통합성 등 ITS만의 독특한 특징을 갖고 있기도 하다. 이런 특성들로 인해 ITS산업은 구(舊) 경제와는 다른 기업전략과 산업정책을 필요로 한다. 네트워크 외부성에 비추어 볼 때, ITS산업에 속하는 기업들은 개방과 공유 또는 전략적 제휴 등을 통해 경쟁사들과 시장을 나누어 가지는 전략이 효과적이며, 시장의 성숙을 위해서는 표준화를 조기에 이룩해야 한다. 자연 독점적 성격과 공공성 및 수익성의 병존이라는 특징에 비추어 ITS산업은 시스템별로 공공성 및 수익성 정도를 검토한 후, 이에 근거하여 공급주체와 자금조달주체를 결정하는 것이 바람직하다. 본 연구는 정보경제에서 ITS산업의 특성과 이러한 특성이 기업전략과 산업정책에 미치는 영향에 대해 분석하여 보았다.