• Korean Journal of Animal Reproduction
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• 제25권2호
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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.

• Reproductive and Developmental Biology
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• 제31권3호
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• pp.169-174
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• 2007

Despite many efforts to improving canine in vitro maturation(IVM), the efficiency is still low compared to that of other mammalian species. The present study investigated the effects of gonadotropin, epidermal growth factor and cysteine supplementation on in vitro maturation of canine oocytes. Cumulus-oocyte complexes (COCs) were matured in basic medium (TCM-199 containing 10% FBS, 0.11mg/ml sodium pyruvate supplemented with or without $10{\mu}g/ml$ FSH, 10ng/ml EGF and 0.57mM cysteine) for 72 hr at $38.5^{\circ}C$ in humidified atmosphere of 5% $CO_2$ in air. After culture, oocytes were stained with Hoechst 33342 $(10{\mu}g/ml)$ for 30 min at $4^{\circ}C$, and assessed their nuclear status (GV: germinal vesicle, GVBD: germinal vesicle break down, MI: metaphasse I, MII: metaphase II, UK: unknown stage). No differences were observed in GV, MI and MII rate except GVBD rate between with and without gonadotropins addition respectively (6.7%, vs. 17.2%). Supplementation of 10ng/ml of EGF in hormones added IVM medium resulted in significantly (p<0.05) higher MII rate (4.54% vs.7.06%). Although there are no significantly difference, total of MI and MII rates were increased by adding cysteine. In conclusion, the present study indicates that supplementation of $10{\mu}g/ml$ LH and FSH, 10ng/ml EGF and 0.57mM cysteine in canine IVM medium show a positive influence on the progression of maturation to MII at 72 hr.

• Journal of the Korea Academia-Industrial cooperation Society
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• 제20권9호
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• pp.333-340
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• 2019

This study investigated the effects of goat blood serum (gBS), goat follicular fluid (gFF) and gonadotropin (FSH) on the in vitro maturation, fertilization and development of Korean native black goat oocytes. Our results indicate that the gBS combined with FSH treated group showed significantly higher maturation rate than the other groups. Furthermore, blastocyst formation rate was significantly increased in all treated groups, and gBS and gFF combined with FSH treated groups were higher than other groups. However, gene expression levels of BMP15 and GDF9 in COC, both oocyte maturation related genes, remained unaffected after 24 h maturation. The results of the present study indicate that supplementation of the maturation medium with gBS, gFF and FSH is efficacious in improving the in vitro maturation, fertilization and development of Korea native black goat oocytes.

• Journal of Embryo Transfer
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• 제19권2호
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• pp.81-87
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• 2004

This study was designed to determine whether repeated superovulation is beneficial for recovery and quality of oocytes in Korean native goats. Seventy-six mature goats, maintained in a pen under natural day length and fed hay ad libitum, were pretreated with progestagen impregnated CIDR for 10 days and then the goats were divided into two groups. One group of the goats received a single intramuscular injection of 1,000 IU PMSG on Day 8 of CIDR insertion. The other group of the goats received twice daily intramuscular injections of a total of 70 mg FSH for 3 days from Day 8 of CIDR. All the gonadotropin treated goats were injected with 10 mg $PGF2{\alpha}on$ Day 8 and 400 IU hCG in the afternoon on Day 10. For oocyte recovery, donor goats were fasted 24 h before operation. Anesthesia was induced by intravenous injection of 2% xylazine(0.2 mg/kg body weight) and ketamin(11 mg/kg body weight). In vivo oocytes were recovered by follicle aspiration or oviduct flushing at 35 to 40 hours after hCG injection through mid-ventral incision. The mean number of CL and oocytes recovered and recovery rate of oocytes by oviduct flushing were greater(P<0.05) in the first treatment than those in the second treatment. Contrary to our assumption, PMSG treatment significantly (P<0.05) increased the number of CL formed and recovery rate of oocytes compared to FSH. However, the same effect was not observed in recovery of follicular oocytes. There was no significant difference in oocyte quality between FSH and PMSG or first and second treatments. The present results indicate that repeated superovulation and repeated use of donor animals may be inefficient for obtaining oocytes in good qualities.

• Development and Reproduction
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• 제13권4호
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• pp.353-362
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• 2009

GnRH and its receptor are known to express locally in the ovary and to regulate the ovarian function by affecting on granulosa and lutein cells. It has been reported that GnRH directly causes apoptosis in the granulosa and lutein cells of the ovary. However, whether the apoptosis of the cells by GnRH is recovered by FSH as an anti-apoptotic factor is not yet known. In this study, we evaluated the apoptosis and the production of progesterone $(P_4)$ and estradiol $(E_2)$ after treatment with 5, 50, and 100 ng/$m\ell$ GnRH and 1 IU/ml FSH in the granulosa-lutein cells that are obtained during oocyte-retrieval for IVF-ET. Results of DNA fragment analysis and TUNEL assay demonstrated that DNA fragmentation and the rate of apoptotic cells were increased in a dose-dependent manner showing a significant increase in the cells treated with 100 ng/$m\ell$ GnRH. In addition, we found that FSH suppresses the apoptosis of the cells induced by GnRH. In the results of chemiluminescence assay for $P_4$ and $E_2$, $P_4$ production was decreased by GnRH treatment, whereas $E_2$ production was not changed. We also demonstrated that FSH inhibits the suppressive effect of GnRH on $P_4$ production as the result of apoptosis. The present results suggest that GnRH agonist using in ovarian hyperstimulation protocol might induce the dysfunction of the ovary, but its function could be recovered by FSH. These results also will be expected to use as the basic data to elucidate the physiological role of GnRH and to develop new ovarian hyperstimulation protocols for IVF-ET.

• Clinical and Experimental Reproductive Medicine
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• 제35권2호
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• pp.155-162
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• 2008

Objectives: The aim of this study was to assess the change of ovarian reserve after removal of ovarian tumor using basal FSH, $E_2$, clomiphene citrate challenge test and ovarian volume. Methods: Twenty two patients with unilateral ovarian tumor, ${\leq}35$ years old, regular menstrual cycle were collected prospectively and divided into endometrioma or non-endometrioma group. We measured the ovarian volume with transvaginal ultrasonography on the day 3 of menstrual cycle within one month before and 3 months after surgery. Basal (cycle day 3) FSH, $E_2$ and CCCT were checked before surgery and repeated at least 2 spontaneous cycles later after surgery. Three patients that had been pregnant within 3 months after surgery were excluded in analysis. Results: The ovarian volume was reduced significantly after surgery in endometrioma and non-endometrioma ${\geq}10\;cm$ group ($4.79{\pm}2.57\;cm^3$ and $5.21{\pm}1.33\;cm^3$, respectively), but not in the non-endometrioma <10 cm group ($6.18{\pm}2.85\;cm^3$). After surgery, basal FSH and cycle day 10 FSH on CCCT in endometrioma and non-endometrioma were $4.25{\pm}0.20\;mIU/ml$ and $3.79{\pm}0.80\;mIU/ml$, $4.24{\pm}0.85\;mIU/ml$ and $4.28{\pm}0.92\;mIU/ml$, respectively. There were neither significant difference in comparison with the preoperative results nor between two groups. Conclusions: Enucleation of ovarian mass was associated with a significant reduction in ovarian volume in endometrioma and non-endometrioma larger than 10cm in diameter. Hormonal markers for evaluation of ovarian reserve, such as basal and cycle day 10 FSH on CCCT, were not changed significantly in each group. In reproductive age women, conservative enucleation or cystectomy rather than oophorectomy should be considered even in a large benign tumor and ovarian function could be reserved by meticulous operative technique.

• Clinical and Experimental Reproductive Medicine
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• 제24권2호
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• pp.217-223
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• 1997

This study was performed to determine if women with day 3 serum inhibin-B concentrations <45pg/ml (conversion factor to SI unit, 1.00) demonstrate a poor response to ovulation induction and assisted reproductive technology outcome to women with inhibin-B${\ge}45pg$/ml, independant of day 3 FSH, E2 and patient age. From Jan 1996 to Dec 1996, 16 volunteers patients who underwent 25 IVF cycles with luteal phase GnRH agonist suppression and HMG stimulation were allocated to the study group. We evaluated day 3 serum inhibin-B, FSH, E2, peak E2, cancellation rate per initiated cycle (%) and clinical pregnancy rate per initiated cycle (%) according to the above two groups and independent of patient age, day 3 FSH, day 3 E2 and all of above combined. Women with day 3 serum inhibin-B${\ge}45pg$/ml demonstrated higher average day 3 inhibin-B level, clinical pregnancy rate per initiated cycle ($20.3{\pm}2.5$ pg/ml vs $80.9{\pm}5.0$ pg/ml, p<0.05; 24.8% vs 8.5%, p<0.05) and lower day 3 FSH level, cancellation rate per initiated cycle ($6.9{\pm}0.3$ mIU/ml vs $8.5{\pm}0.5$ mIU/ml, p<0.05; 1.5% vs 9.0%, p<0.05). Women with day 3 serum inhibin${\ge}45pg$/ml and age<40 year demonstrated higher pregnancy rate per initiated cycle (28.2% vs 7.4%, p<0.05) and lows. day 3 FSH level, cancellation rate per initiated cycle ($6.9{\pm}0.5$ mIU/ml vs $8.2{\pm}0.7$ mIU/ml, p<0.05; 1.0% vs 9.0%, p<0.05). Women with day 3 serum inhibin${\ge}45pg$/ml and day 3 FSH<15mIU/ml demonstrated higher pregnancy rate per initiated cycle (33.5% vs 9.5%, p<0.05) and lower day 3 FSH level, cancellation rate per initiated cycle ($7.7{\pm}0.2$ mIU/ml vs $8.5{\pm}0.5$ mIU/ml, p<0.05; 1.5% vs 10.0%, p<0.05). Women with day 3 serum inhibin${\ge}45pg$/ml and day 3 E2<50pg/ml demonstrated higher pregnancy rate per initiated cycle (30.0% vs 9.5%, p<0.05) and lower cancellation rate per initiated cycle (1.5% vs 9.5%, p<0.05). Women with day 3 serum inhibin${\ge}45pg$/ml, age<40 year, day 3 FSH<15mIU/ml and day 3 E2<50pg/ml demonstrated higher pregnancy rate per initiated cycle (30.0% vs 10.8%, p<0.05) and lower day 3 FSH level, cancellation rate per initiated cycle ($6.8{\pm}0.6$ mIU/ml vs $8.4{\pm}0.9$ mIU/ml, p<0.05; 1.5% vs 7.8%, p<0.05). Therefore women with low day 3 serum inhibin-B concentrations demonstrate a poorer response to ovulation induction and are less likely to conceive a clinical pregnancy though ART relative to women with high day 3 inhibin-B and day 3 serum inhibin-B, in addition to a day 3 FSH, E2 and patient age, appears helpful in prediction in IVF-ET outcome.

• 대한생식의학회:학술대회논문집
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• 대한불임학회 1996년도 추계 학술대회 및 정기총회
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• pp.67.1-67.1
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• 1996

• 대한생식의학회:학술대회논문집
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• 대한불임학회 2004년도 제46차 춘계학술대회
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• pp.82-83
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• 2004

• 대한생식의학회:학술대회논문집
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• 대한불임학회 1997년도 제34차 추계 학술대회
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• pp.45.1-45.1
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• 1997