• Asian-Australasian Journal of Animal Sciences
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• v.14 no.12
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• pp.1675-1677
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• 2001

The effect of the presence or absence of corpus luteum in the ovaries of slaughtered buffaloes was studied for the oocytes recovery and their subsequent maturation and fertilization in vitro. On an average, 0.41 and 0.67 oocytes per ovary were recovered from ovaries with and without corpus luteum, respectively. Immature oocytes were matured in TCM-199 medium. Significant difference was observed in maturation rate between good (74%) and fair (37%) oocytes. However, there was no significant difference in cleavage rate between the two types. The results of this study show that although the presence of corpus luteum in the ovary at the time of recovery significantly affected availability of total oocytes and in-vitro maturation, but fertilization and cleavage remained unaffected under in vitro conditions.

• Journal of Embryo Transfer
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• v.5 no.1
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• pp.21-27
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• 1990

The technique for maturation of follicular oocyte has been devised to provide such a low cost and in ptentifut number supply of bovine embryo. Some of problems concerning production of bovine embtyo in vitro were discussed in this paper. Bovine follicular oocytes cultured in vitro achieved normal fertilization but cleavage rates to blastocyst were low compared to the oocyte matured in vivo. It has been concluded that a deficient cytoplasmic maturation occurs in the oocytes matured in vitro. These results indicate that the studies for maturation of bovine follicular oocytes in vitro need improvement of culture conditions and to define the characteristics that might be indicative of healthy oocyte.

• Korean Journal of Animal Reproduction
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• v.14 no.3
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• pp.223-230
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• 1990

This experiment was carried out to investigate in vitro maturation rate of pig follicular oocytes cultured from 30 to 48hr in TCM 199 supplemented with gonadotropins(FSH, LH) and estradiol-17$\beta$ and in vitro fertilization with ejaculated sperm preincubated in BO medium containing 2mM caffein and development of IVF oocytes. The results obtained in this experiments were as follows ; 1. In addition of hormones, in vitro maturation rate of follicular oocyte increased gradually from 36hr and 74.47% at 48hr in addition of hormones, but there was no differences among in vitro maturation rates after 36hr of culture. 2. Penetration rate of pig oocytes matured in FSH＋LH＋E2 and FSH＋E2 was 71.8%, 71.0% and significantly increased by the addition of hormones. 3. Percentage of developed oocytes was 44.4% for oocytes matured in FSH＋LH＋E2-added medium and 48.7% for oocytes matured in FSH＋E2-added medium, respectively. 4. Two to 16 cells stage embryos were obtained only when pig oocytes matuerd in vitro in hormones-added medium and 72hr after IVF. 5. From present results, it is concluded that gonadotropins and estradiol17$\beta$ can enhance in vitro fertilization and subsequent development as well as in vitro maturation pig follicular oocytes.

• Korean Journal of Animal Reproduction
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• v.23 no.3
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• pp.239-245
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• 1999

The effect of catalase on in-vitro maturation in porcine oocytes with or without cumulus cells were studied. The maturation rates were not significantly different in medium with and without catalase during in vitro maturation of oocytes. However, the maturation rates of oocytes with cumulus cells were significantly higher (P＜0.05) than in oocytes without cumulus cells regardless of the presence of catalase. On the other hand, the maturation rate in oocytes cultured with cumulus cells for 48 h (57%) was significantly (P＜0.05) higher than in oocytes with cumulus cells for first 24 h period (42%) only. In another experiment, the maturation rate was significantly (P＜0.05) higher in medium containing catalase for last 24 h period only than in medium containing catalase for first 24 h period during in vitro maturation of ooytes with or without cumulus cells. But the oocytes matured to M-II stage were observed at 24 h of culture of oocytes without cumulus cells only. When oocytes with cumulus cells were cultured for 72 h, the maturation rates was significantly (P＜0.05) higher in medium with (79%) than without (65%) catalase during in vitro maturation. These results indicate that cumulus cells are necessary for in vitro maturation of porcine oocytes, catalase have effect according to the addition periods and can prevent aging of porcine oocytes during maturation in vitro.

• Journal of Veterinary Clinics
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• v.27 no.1
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• pp.46-49
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• 2010

In this study, we investigated the number of follicles, oocyte recovery rate and oocyte competence after in vitro maturation according to the size of follicle. And equine oocyte competence after in vitro maturation was investigated in terms of the diameter of follicle with criteria of maturation: nuclear stage after Hoechst staining. The average number of follicles per ovary with middle size (11-20 mm, 2.68) was higher than those of small (5-10 mm, 0.74) and large size follicle (> 21 mm, 1.63), therefore medium follicle (53.1%) had higher proportion than other size of follicles. The average numbers of follicle per ovary was 5.05. The rate of oocyte recovery in small (54.5%) and middle follicle (50%) was higher than that in large follicle (40.9%). After culture for 48 h in Medium 199, 50%, 45.5%, and 44.4% of oocytes from the follicles with diameters of 5-10, 11-20, > 21 mm, respectively reached the metaphase II stage. This is the first report showing number of follicle, oocyte recovery rate according to follicular size, and in vitro oocyte maturation in Jeju mare in Korea. To fulfill in vitro equine embryo production, further studies such as the seasonal effect, in vitro fertilization etc is need.

• Reproductive and Developmental Biology
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• v.29 no.4
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• pp.207-212
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• 2005

In the present study, we investigated the effects of genotypes on in vitro maturation and fertilization in porcine fresh/frozen-thawed oocytes. The porcine cumulus-oocyte complexes (COCs) were divided into four groups according to whether they were: (1) in vitro matured; (2) cryopreserved and in vitro matured; (3) in vitro fertilized and (4) cryopreserved, and in vitro fertilized. Maturation of porcine COCs was accomplished by incubation in NCSU23 medium. Immature oocytes were cryopreserved by Open Pulled Straws (OPS) method according to Vajta et al., (1998). Oocytes stained by Acetic-Orcein method were observed under the microscope. DNA extracted from the ovaries was analyzed by RAPD (random amplified polymorphic DNA) and SSCP (single strand conformational polymorphisrrt) method. The rates of oocytes maturation and fertilization were significantly high in AA genotype. The results indicated that in vitro maturation and fertilization in porcine fresh/frozen-thawed oocytes may be affected by genotypes in pigs.

• Korean Journal of Animal Reproduction
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• v.21 no.1
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• pp.53-59
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• 1997

This study was conducted to investigate the effect of hormones on in vitro maturation of porcine follicular oocytes. The results obtained were as follows : 1. The maturation rates of oocytes cultured in medium containing FSH 1, 10, 50 and 100$\mu\textrm{g}$/ml were 44.6, 58.5, 42.6 and 37.9%, respectively. And those were no difference to maturation rates among the medium containing FSH. However, the maturation rate(13.7%) of oocytes cultured without FSH was significantly(P<0.05) lower than those of oocytes cultured with FSH. 2. The maturation rates of oocytes cultured in medium containing hCG 0, 1, 10, 50 and 100IU/ml were 12.2, 11.9, 17.9, 21.9 and 45.6%, respectively. The maturation rate of oocytes cultured with 100IU/ml hCG was significantly(P<0.05) higher than those of oocytes cultured with 0~50IU/ml hCG concentrations. 3. The maturation rates of oocytes cultured in medium containing E2 0, 1, 10, 50 and 100$\mu\textrm{g}$/ml were 13.7, 10.5, 14.9, 11.4 and 5.9%, respectively. There were no differences to maturation rates among the E2 concentrations. 4. The FSH＋hCG treatment was the highest maturation rate in medium containing different combination of FSH, hCG and E2.

• Reproductive and Developmental Biology
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• v.28 no.4
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• pp.235-240
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• 2004

In vitro maturation of denuded porcine immature oocytes can be enhanced by co-incubation with spermatozoa even before fertilization. This study was to determine whether the addition of spermatozoa into the culture medium could influence the nuclear maturation of porcine cumulus-enclosed germinal vesicle (GV) oocytes. Cumulus-oocyte complexes (COCs) were collected from follicles of 3- to 5-mm diameter. Porcine COCs were cultured in tissue culture medium containing spermatozoa. After 48 h culture, oocytes were examined for evidence of GV breakdown, metaphase I, anaphase-telophase I, and metaphase II. The proportion of oocytes reaching at metaphase II was significantly (P < 0.05) increased in the oocytes cultured in media containing spermatozoa compared to those in media without spermatozoa (52.3% vs 12.5%). No difference in the percentage of metaphase II was observed among the different periods of spermatozoa exposure and among the spermatozoa from different species. The proportion of oocytes reaching metaphase II was significantly different between high and low concentrations of spermatozoa. The present study suggests that manunalian spermatozoa contain a substance(s) that improves nuclear in vitro maturation of porcine cumulus-enclosed GV oocytes. Enhancing effect of spermatozoa for in vitro maturation of oocytes is a highly dose-dependent.

• Korean Journal of Animal Reproduction
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• v.19 no.4
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• pp.251-258
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• 1996

These studies were carried out to investigate the effect of gonadotropins (GTH), fetal calf serum (FCS), porcine follicular fluid (pFF) and FCS and pFF fractions obtained by the gel filtration on in vitro maturation of porcine follicular fluid. When the oocytes were cultured in TCM-199, the maturation rate was higher in pFF than in FCS in both with or without GTH and in pFF the maturation rate was higher in with GTH than in without GTH. In case of without GTH, pFF increased maturation rates in TCM-199, but not in Whitten's medium (WM). When the oocytes were cultured in WM supplemented with FCS fractions, the maturation rate(51.6%) of oocytes was significantly (P<0.05) higher in fraction B (about 30∼70 kDa) than in control, FCS and other fractions. When oocytes were cultured in WM supplemented with pFF fractions, fractions B (about 30∼70 kDa) and D (about 1∼10 kDa) were significantly (P<0.05) higher than in control, pFF and other fractions. In conclusiion, the addition of gonadotropins into the maturation media was effective for oocyte maturation. The addition of pFF was more effective than addition of FCS for maturation of porcine oocytes in vitro. And fraction B from FCS and fractions B and D from pFF was effective for oocyte maturation.

• Journal of Embryo Transfer
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• v.12 no.3
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• pp.293-300
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• 1997

Supplementation of maturation medium with additional granulosa cells has beneficial effect on in vitro maturation of bovine follicular oocytes and their subsequent cleavage and development in vitro. However, maturation system using granulosa cells have some disadvantages that collection of granulosa cells is cumbersome and metabolic activity of the cells is variable according to ovarian cycle or follicular size. We hypothesized that bovine immsture oocytes matured without granulosa cell coculture can fertilize and develop normally if the medium volume per oocyte is reduced during in vitro maturation. Immature oocytes were matured for 24 hours in a TCM199 containing 10% fetal calf serum, anterior pitultary hormone (0.02 AU /ml Antrinⓡ) and estradiol with or without granulosa cells in vitro. In Group 1, 35 to 40 oocytes were matured in a well of 4-well plastic dish containing 500 $\mu$l of maturation medium and granulosa cells, and 9 to 10 oocytes were matured in a 50-$\mu$l drop of maturation medium without granulosa cells in Group 2. After maturation, oocytes were coincubated with sperm for 30 hours in a modified Tyrode's medium (IVF). Inseminated oocytes were cultured in a microdrop (30 $\mu$l) of a synthetic oviduct fluld medium (SOFM) containing BSA, Minimum Essential Medium essential and non-essential amino acids for 9 days. As a preliminary experiment, we investigated the beneficial effect of granulosa cells during maturation on subsequent cleavage and development using the same type of culturedishes (4-well dish). Granulosa cells could not increase embryo cleavage after fertilization but significantly improved (p<0.05) embryo development to expanding blastocyst (Table1 and 2). In Group 1, 68 and 80% of inseminated oocytes have cleaved at 30 hours and 2 days after IVF, respectively, which is similar (p>0.05) to the result of Group 2 (69% at 30 hours and 78% at 2 days after IVF). The oocytes in Group 2 showed 21 and 11% of developmental rates to expanding and hatching blastocysts, respectively, which was not significantly different (p>0.05) from those (20 and 10%, respectively) of oocytes in Group 1. In conclusion, it has been clarified that a microdrop culture system without granulosa cells for in vitro maturation can support bovine embryonic development to blastocyst in vitro as readily as a granulosa cell coculture system.