• 한국응용곤충학회지
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• 제52권4호
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• pp.295-304
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• 2013

파밤나방(Spodoptera exigua)의 발육을 일으키는 최저온도를 결정하고, 이 상태의 생리적 특성을 서로 다른 기능군(대사, 신경, 면역 및 스트레스) 유전자의 발현 양상을 이해하기 위해 본 연구를 수행하였다. 알부터 번데기까지 파밤나방의 발육영점온도는 $5.5{\sim}11.6^{\circ}C$로 다양하였다. 유충은 알과 번데기에 비해 비교적 낮은 온도에서 발육이 가능하였다. 5령충의 경우 생리적 발육영점온도가 추정치($10.3^{\circ}C$)와 다르게 이보다 높은 $15^{\circ}C$에서 관찰되었다. 정량적 RT-PCR로 분석된 유전자의 발현양상은 유충 영기가 진행됨에 따라 모든 기능군의 대부분 유전자의 발현량이 증가하였고, 또한 5령 시기에서도 처리온도가 증가함에 따라 이들 유전자의 발현량도 증가하였다. 비록 동일한 갓 탈피한 5령이라 하더라도 이전에 노출된 외부 온도에 따라 발현량이 상이하였다. 5령충의 생리적 발육영점온도인 $15^{\circ}C$에서 대부분의 유전자 발현량은 저하되었다. 그러나 높은 온도에서와 마찬가지로 발육기간이 증가함에 따라 이들 유전자의 발현량이 증가하였다. 이상의 결과는 발육영점온도에서 파밤나방의 발육 관련 유전자의 발현이 전체적으로 수준은 낮지만 지속적으로 진행되고 있다는 것을 의미한다.

• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 2003년도 제3회 국제심포지움 및 학술대회
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• pp.123-123
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• 2003

This study was conducted to determine the ability of nuclear development of canine oocytes depend on the kind of maturation media and addition of serum sources. Ovaries were collected from a bitches at various stages of estrus cycle by an ovariohysterectomy. Oocytes were collected of cumulus oocytes complexes after slicing of ovaries with blade. The maturation medium was containing 0.6 mM/ml cysteine, 0.2 mM pyruvic acid, 20 ng/ml $E_2$ and 1 $\mu g/ml$ rbST Exp. 1, the oocytes were matured in four different maturation medium as follows: 1) TCM-199, 2) DMEM, 3) NCSU37 and 4) modified-NCSU37 with 10% FBS. Exp. 2: the oocytes were matured in mNCSU37 supplemented with different protein sources (10% FBS, 10% EDS, 0.3% BSA and 0.1% PVA) to select the optimal one. Oocytes were matured in a humidified atmosphere containing 5% $CO_2$ at $39{\circ}C$ for 72 hrs. The maturation rate were analyzed by Duncan's multiple range test using General Linear Models procedure in SAS. The rates of meiotic resumption to MI-MII depend on different culture media were achieved with TCM-199 (5.2%), DMEM (5.0%), NCSU37 (7.2%) and m-NCSU37 (5.9%), respectively. The rates of meiotic resumption to MI-MII according to addition of protein source were 10% FBS (13.3%), 10% EDS (25.0%), 0.3% BSA (25.0%) and 0.1% PVA (15.4%), respectively. In conclusion, the results obtained showed that in vitro maturation media and protein supplement to m-NCSU37 culture medium tested did not promote the final steps of IVM in canine oocytes.

• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 2003년도 제3회 국제심포지움 및 학술대회
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• pp.122-122
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• 2003

This study examined the viability of canine oocytes following storage at 4 or $38{\circ}C$ for 5 hrs. The ovaries were collected from domestic dog following ovariohysterectomy at a local veterinary clinics and transported to laboratory In two different transport temperature at 4 or $38{\circ}C$ within 5 hrs. The cumulus oocyte complexes (COCs) were recovered after slicing with blade. In Exp. 1, the oocytes collected were matured in DMEM supplemented with l0% FBS, 0.6 mM/mlcysteine, 0.2 mM Pyruvic acid, 20 ng/ml $E_2$ and 1 $\mu g/ml$ rbST at humidified atmosphere containing 5% $CO_2 38{\circ}C$ for 24 or 48 hrs to analysis of survivability. In Exp 2, to assess nuclear development at 38{\circ}C$ group, the oocytes were matured in maturation medium for 24, 48 or 96 hrs. Survivability was judged by a morphological appearance and PI staining. Survivability rates were analyzed by General Linear Models procedure in SAS. The survival rates at 4{\circ}C$ ovary transport group showed significantly lower than at 38{\circ}C$ group (0 vs 72.9% in 48 hrs and 13.2 vs 77 8% in 24 hrs; P<0.05). The nuclear development of oocytes to MI to MII stages at 24, 48 and 96 hrs was 8.3% (6/72), 8.9% (9/101), and 9.5% (8/84). These results showed that the canine oocytes were remarkably sensitive to a low temperature and did not increase nuclear development rate depend on maturation time to 96 hrs.

• 한국발생생물학회:학술대회논문집
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• 한국발생생물학회 2003년도 제3회 국제심포지움 및 학술대회
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• pp.74-74
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• 2003

In present study, attempts were made to preserve abalone (Haliotis discus hannai) sperm in liquid form at low temperature, to evaluate the effect of various diluents in short-term storage on sperm, and cryopreservation procedures were optimized for the cryoprotectants as well as freezing rates, in terms of the motility and survival rate, and the ultrastructural changes of sperm after short-term storage and cryopreservation were observed. The abalone sperm reached maximum motility until about 4min after activation. The motility was constant for about 16min, after which it dropped gradually, and about 50min later all motility ceased. Threshold activation of sperm was found in 40% artificial seawater (ASW), and motility increased as the concentration of ASW increased. In Hanks balanced salt solution without calcium (Ca-Free HBSS, 300 and 400 mOsmol/kg) and 10%, 20%, and 30% ASW the sperm was immotile, and motility once again restored incompletely only in HBSS of 300 and 400 mOsmol/kg, 20% and 30% ASW after 100% ASW was added. Sperm motility was extended following 20 days of cold storage only in 70% and 100% ASW. A high motility index of 3.5-4.5 was observed for the first 8 days in 70% and 80% ASW. In other diluents sperm motility was constant less than 10 days, and the motility index was obviously lower than that of sperm in 70% and 100% ASW. After 20 days of cold storage survival rates of 10.2%-20.7% were obtained in ASW and 300 mOsmol/kg HBSS, and that in 400 HBSS (65.3%) was significantly higher than others. The constant period of sperm motility stored in 70% ASW was longer obviously than that in 100% ASW after 6 days of storage, and the time to maximum motility of sperm stored in 70% increased gradually, while the difference in which of sperm in 100% ASW was not significant. The sperm plunged into liquid nitrogen all died except that sperm using 15% glycerol as cryoprotectant restored 10.4% of motility. The highest motility index (3.4) was obtained with 5% glycerol and freezing procedure: $50^{\circ}C$/min from $20^{\circ}C$ to $-80^{\circ}C$.

• Clinical and Experimental Reproductive Medicine
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• 제28권2호
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• pp.161-167
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• 2001

Objective: Granulocyte-macrophage colony stimulating factors known to be secreted in murine and human reproductive tract. The development of human, bovine and murine embryos could be promoted by addition of GM-CSF in culture medium. However, the pregnancy and implantation rate of embryos cultured in GM-CSF have not been evaluated. The aim of this study was to assess the effect of GM-CSF in embryo development, pregnancy and implantation rate. Methods: A total of 191 IVF cycles were divided into control and GM-CSF supplement group (control=96, GM-CSF=95). The embryos were cultured for three day with or without 2 ng/ml of recombinant human GM-CSF. The quality of embryo, developmental velocity, pregnancy and implantation rates were compared. Results: There was no difference in age, number of gonadotropin ampules used, number of oocytes and fertilization. The number of ICSI cycle was higher in GM-CSF group. In GM-CSF group, G-1 grade embryos were the highest in proportion (56.4%), while G-2 grade embryos were highest (44.3%) in control group. The developmental velocity of embryos were not different between GM-CSF and control group. The pregnancy and implantation rates were significantly higher in GM-CSF group than control (47.4% vs. 33.3%, 17.0% vs. 11.1% respectively). Conclusion: By adding GM-CSF in culture medium, the quality of embryo, pregnancy and implantation rate could be improved.

• 한국수정란이식학회지
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• 제10권2호
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• pp.131-138
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• 1995

Essential and non-essential amino acids supplemented to culture medium stimulate mammalian embryo development in vitro. Amino acids such as glycine, taurine and alanine are concentrated in the lumen of oviduct and uterus and it can he thought that these amino acids may have physiological role on fertilization and embryo development. Our aim of this experiment was to investigate the effects of essential and non-essential amino acids, taurine or glycine supplemented to fertilization medium on the cleavage and subsequent in vitro development of bovine oocytes matured and fertilized in vitro. Immature oocytes were obtained from slaughtered Holstein cows and heifers and matured in TCM199 containing 10% fetal calf serum, 2.5 $\mu$g /mL of FSH and LH and 1 $\mu$g / mL of estradiol with granulosa cells in vitro. After maturation, oocytes were coincubated with sperm in fertilization medium supplemented with Minimum Essential Medium (MEM) essential and non-essential amino acids, taurine (3.75 mM) or glycine (10 mM) for 30 hours in vitro. Inseminated oocytes were cultured in synthetic oviduct fluid medium (SOEM) containing MEM essential, non-essential amino acids and 1 mM glutarnine up to 8 days after fertilization.Supplementation of fertilization medium with MEM essential and non-essential amino acids lowered significantly (p<0.05 and p<0.001) the cleavage rate after 30 hours of IVF (53.3%) and at Day 3 (62.7%: Day 0: the day of I VF) compared to control (64.3% and 77.3%, respectively). Subsequent developmental rates to morulae (Mo) and expanding blastocysts (ExBL) also significantly decreased (p<0.001 and p<0.05 for Mo and ExBL) when oocytes were coincubated with sperm in the medium containing MEM amino acids. Taurine added to fertilization medium have not increased the cleavage rate over the control, whereas glycine showed significantly lower (p<0.01) cleavage rate at Day 3 than that of taurine, but there was no significant difference in the developmental rates to Mo and ExBL of bovine embryos irrespective of the supplementation of taurine or glycine to fertilization medium. In conclusion, supplementation of fertilization medium with essential and non-essential amino acids, taurine or glycine has no beneficial effect on in vitro cleavage and development of bovine oocytes matured and fertilization in vitro.

• 한국수정란이식학회지
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• 제21권3호
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• pp.183-190
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• 2006

본 연구는 고급육을 생산한 한우의 유전자를 재활용하기 위해 기초 등록우와 혈통 등록우의 육질등급이 1 등급 이상인 암소의 난소에서 개체별로 채취한 난포란을 개체별로 각각 체외 성숙, 체외 수정 및 체외 배양하였다. 그 결과를 요약하면 다음과 같다. 기초 등록우의 육질 등급별로 회수된 난포란은 각각 1 등급 평균 28.9개, $1^+$등급 평균 28.8개, $1^{++}$ 등급 29.6개로 평균 29.0개였다. 기초 등록우의 육질 등급별 배반포배 형성율은 각각 1 등급 27.2%, $1^+$ 등급 28.7%, $1^{++}$ 등급 32.9%로 평균 8.4개의 배반포배가 생산되었으며, 배반포배 형성율은 육질등급간에 유의적 차이가 없었다. 혈통 등록우의 육질 등급별로 회수된 난포란은 각각 1 등급 평균 25.8개, $1^+$등급 평균 27.1개, $1^{++}$ 등급 27.0개로 평균 26.7개였다. 혈통 등록우의 육질 등급별 배반포배 형성율은 각각 1 등급 23.0%, $1^+$ 등급 33.7%, $1^{++}$등급 42.6%로 평균 8.6개의 배반포배가 생산되었으며, 배반포배 형성율은 육질등급간에 유의적 차이가 있었다(p<0.05).

• Clinical and Experimental Reproductive Medicine
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• 제25권2호
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• pp.129-134
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• 1998

본 연구는 인간 전핵기 수정란의 동결시 election microscope grid론 사용하는 초급속 동결방법이 유용하는지 여부를 조사하고자 실시하였다. 본 실험에서는 인간 IVF 시술시 생성되는 다-전핵기 (>2PN) 수정란을 정상 2PN 수정란 대신 사용하였으며, 또한 이들은 3PN과 $\geq4PN$ 수정란으로 나누어 전핵의 수에 따른 냉해와 융해 후 체외 배발달율을 조사하였다. 동해제는 30% ethylene glycol, 18% ficoll, 0.5 M sucrose와 10% FBS 등이 D-PBS에 첨가되어 제작된 EFS30을 사용하였다. 본 연구에서 얻어진 결과는 다음과 같다. 초급속동결-융해 후, 인간 다-전핵기 수정란의 생존율은 85.5%였다. 대조군과 동결군의 난할율을 진핵의 수에 따가 니누어 비교하였을 때, 각 군간에는 유의한 차이를 나타내지 않았다 (3PN; 81.3%와 85.4%, $\geq4PN$; 90.0%와 95.7%). 또한, 융해 후 체외발달율을 조사하였던바, $\geq4PN$수정란에서는 동결군 (4.5%)의 체외발달이 대조군 (44.4%)보다 유의하게 낮게 나타났던 반면, 3PN 수정란에서는 동결군 (22.0%)의 결과가 대조군 (38.5%)에 비해 유의한 차이를 나타내지 않았다. (p<0.05). 따라서 인간 다-전핵기 수정란은 EM grid와 EFS30 동결액을 이용한 초급속 동결로 배발달을 유도할 수 있다는 것을 알 수 있었다.

• Clinical and Experimental Reproductive Medicine
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• 제40권3호
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• pp.122-125
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• 2013

Objective: The majority of embryo transfers (ETs) to date have been performed on day 3 to reduce the potential risk of developmental arrest of in vitro cultured embryos before ET. Development of sequential media has significantly improved culture conditions and allowed blastocyst transfer on day 5. While day 5 ET provides higher clinical pregnancy outcomes with reduced risks of multiple pregnancies, it still has potential risks of developmental arrest of IVF embryos. The aim of this study was to evaluate the clinical outcomes of day 4 ETs and compare the efficacy of day 4 ET with day 5 ET. Methods: From 2006 to 2009, a total of 747 fresh IVF-ET cycles were retrospectively analyzed (day 4, n=440 or and day 5, n=307). The cycles with any genetic factors were excluded. The rates of matured oocytes, fertilization, good embryos, and clinical pregnancy of the two groups were compared. The chi-square test and t-test were used for statistical analysis. Results: There were no significant differences between the two groups with respect to the mean age of the females and rates of matured oocytes. The pregnancy outcomes of day 4 ET (40.7%) were similar to those of day 5 ET (44.6%). The implantation rate of day 5 ET (24.2%) was significantly higher than that of day 4 ET (18.4%) (p=0.003). Conclusion: Day 4 ET can be chosen to avoid ET cancellation in day 5 ET resulting from suboptimal circumstances in the IVF laboratory, but the decremented quality of embryos for transfer and the decreased pregnancy rate must be taken into consideration.

• 생명과학회지
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• 제23권7호
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• pp.941-946
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• 2013

신경세포의 가지돌기 내 단백질합성은 필요한 단백질을 실시간으로 제공할 수 있는 이점을 제공한다. 본 연구에서는 단백질합성인자 eIF4E와 그 억제 단백질인 eIF4EBP1의 발생단계별 표현을 배양한 해마신경세포를 면역 염색하여 조사하였다. eIF4E는 가지돌기에 점박이 모양으로 표현되었으며, 핵에는 표현되지 않았다. 그러나 eIF4EBP1는 가지돌기 뿐 아니라 발생초기(DIV 0.5)부터 핵에서 표현되었으며 성숙한 세포에서 핵에 더욱 뚜렷이 표현되었다. eIF4E 혹은 eIF4EBP1의 PSD95과의 colocalization은 $39.1{\pm}9.6%$ 및 $70.5{\pm}5.2%$ (DIV 7), $57.7{\pm}8.2%$ 및 $36.0{\pm}3.1%$ (DIV 10), $29.9{\pm}2.9%$ 및 $40.2{\pm}11.7%$ (DIV 20)이었다. eIF4E와 eIF4EBP1의 colocalizatin은 $18.5{\pm}2.6%$ (DIV 7), $11.1{\pm}3.9%$ (DIV 10), $38.6{\pm}5.6%$ (DIV 20)이었다. 이 결과는 eIF4E 및 eIF4EBP1의 많은 부분이 연접후에 위치하며, 발생초기에는 eIF4E가 활동적인 형태로 존재하지만, 성숙 신경세포에서는 eIF4EBP1과 결합하여 비활성적인 형태로 존재함을 의미한다.