• Korean Journal of Animal Reproduction
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• v.22 no.4
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• pp.319-329
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• 1998

These experiments were conducted to investigate the optimal maturation stage for cryopreservation of porcine oocyte when the oocytes were frozen-thawed and/or exposed in cryoprotect ant at immature, maturing and mature stages. The results of this research are as follows ; 1. When the oocytes matured for 0, 24 and 44h were exposed in media containing cryoprotectants or without in vitro, the rates of cultured oocytes developed to metaphase II were 44.0, 45.0, 50.3 or 55.0%, respectively. 2. When the oocytes matured for 0, 24 and 44h were exposed in media containing cryoprotectants or without in vitro, the cleavage rates of cultured oocytes were 18.6, 19.7, 47.6 or 50.9%, respectively. 3. When the oocytes matured for 0, 24 and 44h were frozen and thawed using vitrification or not in vitro, the rates of cultured oocytes developed to metaphase II were 4.3, 7.1, 46.7 or 62.4%, respectively. 4. When the oocytes matured for 0, 24 and 44h were frozen and thawed using vitrification or not in vitro, the cleavage rates of cultured oocytes were 2.5, 2.4, 10.2 or 49.6%, respectively.

• Reproductive and Developmental Biology
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• v.28 no.3
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• pp.173-179
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• 2004

This study was conducted to investigate the toxi-cological effects of different vitrification solution on development of immature porcine oocytes in vitro. Oocytes were exposed to EFS solution [40% ethylene glycol (EG) + 18% Ficoll + 0.3M sucrose], ES solution (5.5M EG + 1.0M sucrose) or GE solution [10% glycol (G) + 20% EG], and these oocytes were transferred to sucrose solution directly. Maturation rates were significantly (P<0.05) higher in the ES solution (44.5%) and control (57.6%) than in the EFS solution (38.8%) and GE solution (22.4%). No differences among three solution were found in fertilization rates. Cleavage rates was significantly (P<0.05) higher in the ES solution (47.1%) and control (65.9%) than in the EFS solution (21.9%) and GE solution (19.0%), but no difference among three solutions was found in the blastocyst formation rates. These results indicate that combination of EG and sucrose solutions had effects on development of immature porcine oocytes.

• Proceedings of the KSAR Conference
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• 2001.03a
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• pp.34-34
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• 2001

본 연구의 목적은 한우 난포발달에 있어서 난포액 및 anti-inhibin serum의 생리적 역할을 검토하기 위해 수행하였다. Anti-inhibin serum(AI)은 항원으로서 porcine inhibin-$\alpha$-subunit 19~32의 peptide를 사용하여 adjuvant 용액을 1:3의 비율로 혼합하여 앙고라종 토끼 5두(체중 2.5kg)에게 주 2회 간격으로 접종 후 얻어진 항혈청을 사용하였다. 난포액(bFF; bovine follicular fluid)은 도축장에서 도축되는 한우 난소로부터 직경 1.0cm 이하의 난포로부터 회수하여 스테로이드를 제거하기 위해 10% chacoal so lution(50 mg/$m\ell$, Norrit-A, Fisher Sci., USA)을 처리하여 45분간 배양후 원심분리후 상층액을 회수하여 실험에 공시하였다. 공시동물은 1산후 정상적으로 발정주기가 반복되는 한우암소 9두를 난소감정후 황체를 확인하여 PGF2$\alpha$제제(lutylase. USA)를 주사하여 발정을 유기한 다음, 난소의 first wave가 시작되는 시기인 배란직후 12시간째부터 4일간 일일 2회 5 $m\ell$씩 총 8회 40 $m\ell$의 AI와 bFF를 각각 경정맥으로 주사하였으며 대조구로서 생리식염수를 주사하였으며 채혈 및 정맥주사를 용이하게 하기 위해 경정맥에 카테타를 설치하여 6시간간격으로 총 200시간까지 채혈하였으며 초음파진단기를 이용하여 난포의 발달을 검토하였다. 채혈후 혈중Inhibin, progesterone 및 Estrad iol-17$\beta$농도의 분석은 RIA 및 ELISA법으로 분석하여 얻어진 결과는 다음과 같다. 혈중 Progesterone농도는 대조구와 AI처리구에서는 배란후 68시간째부터 유의적으로 증가하기 시작하였으나, bFF처리구에서는 배란후 68시간부터 170시간까지 대조구에 비해 유의적으로 낮은 농도를 나타내었다. 이에 반해 혈중 Estradiol-17$\beta$농도는 대조구의 경우 bFF처리구와 비슷한 수준으로 배란후부터 낮은 농도를 유지하였으나, AI 처리구는 배란후 36시간이후부터 108시간까지 유의적으로 높은 수준을 유지하였다가 그 이후 감소되었다. 한편, 혈중 Inhibin농도는 전 구간에서 배란후 84시간까지 불규칙한 농도를 보이다가 bFF처리구에서는 배란후 84시간부터 유의적으로 증가하였다. 배란후 72시간째에 초음파진단기를 이용하여 난소의 난포발달을 조사한 결과 , 대조구와 bFF처리구에 비해 AI처리구에서 발달난포가 유의적으로 많은 것을 확인하였다. 이상과 같은 결과로, Anti-inhibin serum은 한우 자체에서 분비하는 Inhibin을 특이하게 억제하여 Inhibin에 의해 억제되는 FSH분비가 촉진됨으로써 난포발달과 estrogen의 농도가 촉진되는 것으로 사료되어 anti-inhibin serum이 한우의 과배란유기 효과가 있는 것으로 사료된다.

• Korean Journal of Plant Resources
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• v.14 no.1
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• pp.8-14
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• 2001

The effects of media, cholesterol, $\beta$-sitosterol and lecithin on the growth and oospore production of the isolates KM10, U6, CDB6, MHB6, JD1 (A$^2$type) of Phytophthora infestans isolated in Korea and F8l7, DNC303 (A$^1$type), IB908, DN107 (A$^2$type) obtained from Japan were investigated. Mycelium of P. infestans grew better on V-8 juice agar and rye meal agar than on the other media. Oospores were produced most abundantly on V-8 juice agar. Mycelium extended more 16.6, 8.3, and 5.2% on V-8 juice agar supplemented with 5 $\mu\textrm{g}$/$m\ell$ of cholesterol, $\beta$-sitosterol and lecithin, respectively, and oospores are produced 76.0, 58.0, and 34.6 % on V-8juice agar supplemented with 5 $\mu\textrm{g}$/$m\ell$ of cholesterol, $\beta$-sitosterol and lecithin, respectively. Oospores more produced on detached potato plant disks when $A^1$ and $A^2$ type exist simultaneously which indicating that variation of population can occur in the field, but the rate of oospore formation and the number of oospores produced was low and small quantity.

• Journal of Embryo Transfer
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• v.22 no.1
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• pp.33-37
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• 2007

A 2 years old female beagle dog was examined because of the vaginal discharge and vaginal swelling at 7th day after mating. Abdominal ultrasonography revealed multiple follicular cysts on both ovaries. Also, the ratio of superficial cells was above 90% based on vaginal cytology. She was still in standing heat, with the tail deflected up and flagging when a male sniffed the vulva and attempted to mount. The dog was diagnosed with multiple follicular cysts by vaginal cytology, clinical signs and ultrasonography. The dog was treated with GnRH $50{\mu}g$ injection by intramuscularly. The sizes of the cysts were decreased two weeks after GnRH treatment, and then the dog delivered an offspring at 68th day after last mating. To confirm the stage of estrous cycle, we performed hormone analyses, retrospectively $Estradiol-17{\beta}$ concentration was 78pg/ml at diagnosis. Progesterone concentration was 6.9ng/ml at the last mating, and 66.9ng/ml at diagnosis. Therefore, we confirmed that follicular cysts was observed during diestrous stage in a beagle dog.

• Korean Journal of Animal Reproduction
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• v.25 no.2
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• pp.101-111
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• 2001

The present study was conducted to develop an in vitro culture system that would support bovine follicle growth from preantral to antral stage, oocyte maturation, fertilization, and embryonic development. Bovine preantral follicles (150$\pm$1.2 ${\mu}{\textrm}{m}$) surrounded by theca cell were isolated ezymatically and mechanically from ovarian cortical slides in Leibovitz L-l5 medium containing 1 mg/$m\ell$ collagenase and 0.2 mg/$m\ell$ DNase I and cultured for 25 days in the presence of different concentrations of bovine FSH and LH in $\alpha$MEM medium. The survival and growth rates of follicles cultured in the presence of FSH (10~150 ng/$m\ell$) were significantly higher than those of control group (P ＜ 0.001), but no significant differences were observed in survival and growth rates of follicles between the LH treatment groups (1~125 ng/$m\ell$) and the control. The survival (40%) and growth (244 $\pm$ 0.5 $\mu\textrm{g}$) of follicles cultured with FSH (90 ng/$m\ell$) and LH (25 ng/$m\ell$) were higher than those of control (25%, 160 $\pm$1.0 $\mu\textrm{g}$). Finally, 50% percent of healthy antral follicles were obtained, and almost 60% of them has complete meiotic division with 1st polar body (18.1%) and 10.0% have developed to the cleaved embryo and blastocyst stage. These results suggest that bovine preantral follicle with intact theca cell can grow to the antral stage using these culture conditions, and that oocytes from in vitro-matured bovine preantral follicle may acquire meiotic competence and can undergo fertilization and development.

• Korean Journal of Agricultural Science
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• v.34 no.1
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• pp.13-18
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• 2007

This study was carried out to investigate the effects of ovarian estrus stage of oocytes and supplementation of medium with BSA, cysteine and myoinositol on in vitro maturation of canine oocytes. The in vitro maturation(IVM) rate to metaphase II (M II) stage of oocytes recovered from different stage of the reproductive cycle (inactive, follicular and luteal stage) cultured in TCM-199 media were $0.0{\pm}0.0%$, $10.0{\pm}4.1%$ and $5.7{\pm}1.6%$, respectively. The IVM rate of oocytes collected from follicular stage was significantly higher in inactive or luteal stage of oocytes. The IVM rates of oocytes recovered from different stage of the reproductive cycle cultured in TCM-199 media with 5% BSA and 0.1 mM cysteine were$0.0{\pm}0.0%$, $15.8{\pm}4.7%$, $5.6{\pm}1.5%$, respectively. The IVM rates of oocytes recovered from different stage of the reproductive cycle cultured in TCM-199 media with 5% BSA and 10 mM myoinositol were $0.0{\pm}0.0%$, $18.4{\pm}4.6%$ and $5.7{\pm}1.9%$, respectively. The IVM rate of follicular stage oocytes was significantly higher in oocytes collected from follicular stage and with cultured 5% BSA and 0.1 mM cysteine or 5% BSA and 10 mM myoinositol compared to other experimental group.

• Development and Reproduction
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• v.4 no.2
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• pp.181-186
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• 2000

The expression of glucosamine-6-phosphate deaminase (GNPDA) was examined in mouse ovary from neonate to aduit. In western blot, band of Mr. 31 kDa antigen sharply increased 2 weeks after birth onward. In irmmunostaining of the adult ovary, GNPDA expression was constitutive in the theca and interstitial cells. However, expression in the granulosa cells was different according to folliculogenesis. Cytoplasm of the oocyte of some primary follicle showed positive signal but not in the antral follicle. Granulosa cells of antral follicles showed no visible sign of GNPDA expression. In the corpora lutea, the signal intensity in granulosaluteal cells increased according to luteal development and became the highest in the luteolytic phase. In summary, the differential expression of GNPDA was found in follicle cells according to folliculogenesis. It suggests that GNPDA might be involved in tissue remodeling in mouse ovary.

• Journal of Embryo Transfer
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• v.9 no.1
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• pp.103-110
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• 1994

소 난포란의 체외성숙율과 체외수정후 분할율을 향상시키기 위하여 성숙배양액에 FCS 첨가 효과를 조사하였으며 FCS의 첨가수준은 5-20%였고 사용된 기본배양액은 Dn(-BSA), Ham's F10 및 TCM199이었다. 난포직경 2-6mm 난포로부터 채란된 수정란을 39$^{\circ}C$ 배양기에서 28시간 성숙시킨후 동결융해정자 또는 비동결정자로 체외수정시켰다. FCS 무첨가구보다는 첨가구 성숙유리 향상되었으나 FCS 첨가의DM(-BSA)와 Ham's F10간에, 그리고 FCS첨가수준간에는 성숙율의 차이가 없었다. FCS의 첨가로 성숙율은 향상되었으나 난할율에는 영향이 없었다. 체외수정 난포란의 난할율이 HIS 처리후 6시간 전배양한 동결정자와의 수정에서는 현저히 낮았으나 2시간 전배양과 정자에 의해서는 향상되었고 비동결정자이용시 더욱 향상되었다. FCS 첨가된 Ham's F10과 TCM199에서 성숙시킨 난자의 난할율이 DM(-BSA)+FCS보다 높았다. 본 연구 결과에서 난포란의 체외성숙이 FCS첨가로 개선되었으나 난할율에는 영향이 없었으며 난할율이 정자처리방법에 따라 차이가 많았고 Ham's F10과 TCM199에 FCS의 첨가가 보다 효과적이었다.

• Korean Journal of Animal Reproduction
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• v.25 no.2
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• pp.119-124
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• 2001

This study was performed to investigate the effects of the peptide to carrier ratio on the immune and biological functions to inhibin immunization in Hanwoo. A peptide sequence kom the alpha -subunit (19~32 peptide) of porcine inhibin was synthesized for antigen and conjugated to human serum albumin(HSA) for carrier protein. Anti-inhibin sera(AI) were produced 52 day later from rabbit after injection of inhibin-$\alpha$ -subunit peptide conjugator for antigen with the interval of 2 weeks. Immune-blotting analysis using antibody specific fur inhibin-$\alpha$ subunits revealed that the inhibin was detected at 1.0 cm bovine follicular fluid(bFF). However, each stage of corpus lutea and 0.1 cm of follicular fluid were not detected. The maximal contents of estradiol-17 $\beta$ in Hanwoo ovarian follicular fluid were detected at 2.0 cm of follicular size(diameter), but the mean total contents of these hormone decreased significantly with decreasing diameter of follicles. However, progesterone contents of follicular fluid were high at 1.0 cm of follicle. Progesterone secretion by Hanwoo granulosa cell cultured for 48 hr in vitro was significantly (p＜0.05) inhibited in 5% bFF and 5% bFF + 5% AI addition group compared with control group. Estradiol-17 $\beta$ secretion by Hanwoo granulosa cell cultured for 48 hr in vitro was significantly (p＜0.05) increased in 5% AI and 5% AI + 5% bFF addtion group compared with control group. However, the groups added 5% AI were not changed compared to control groups in progesterone and estradiol-17 $\beta$. Taken together, we suggested that inhibin in the mature FF plays a pivotal role on the biosynthesis of steroid hormone of follicular cells during follicular development.