• 한국수정란이식학회지
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• 제27권2호
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• pp.71-80
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• 2012

Biotechnologies for cloning animals and in vitro embryo production have the potential to produce biomedical models for various researches. Especially, pigs are a suitable model for xenotransplantation, transgenic production and various areas of reproductive research due to its physiological similarities to human. However, utilization of in vitro-produced embryos for transfer remains limited. Despite improvement over past few decades, obstacles associated with the production of good quality embryos in vitro still exist which limit the efficiency of cloning. One of major problems includes improper in vitro maturation (IVM) and culture (IVC). Oxidative stress caused from in vitro culture conditions contributes to inadequate IVM and IVC which leads to poor developmental competence of oocytes, failure of fertilization and embryo development. To reduce the oxidative stress, various antioxidants have been used to IVM and IVC system. However, limited information is available on the effects of resveratrol on livestock reproductions. Resveratrol is a polyphenolic natural product and well known as an antioxidant in foods and beverages (e.g. in grapes and red wine). Resveratrol is known to be cardioprotective, anticarcinogenic, anti-inflammatory, antioxidant and antiapoptotic. This paper will review the effects of resveratrol on in vitro maturation of oocytes and embryo development.

• Reproductive and Developmental Biology
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• 제32권2호
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• pp.71-79
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• 2008

Surface-enhanced laser desorption and ionization time-of-flight mass spectrometry (SELDI-TOF MS) is one of the recently developed proteomic technologies which is based on capturing proteins and peptides by chemically modified surfaces and highly sensitive for the analysis of complex biological samples. In the present study, to gain insights into oocyte maturation and early embryo development, SELDI-TOF-MS was used to find the protein candidates that are specifically or prominently expressed in porcine oocytes at the in vitro matured metaphase II (MIIl) and germinal vesicle (GV) stages. By selected CM10 chip, 16 candidates were found to be up-regulated in GV stage oocytes compared with in MII stage oocytes, their molecular weights were 8,180 (2 candidates), 10,226 (5 candidates), 15,767 (5 candidates) and 16,770 (4 candidates) Da respectively. And the expression of 29 candidates were higher in MII than in GV stage oocytes, their molecular weight were 10,832 (3 candidates), 17,743 (8 candidates), 20,122 (3 candidates), 22,131 (3 candidates), 24,857 (7 candidates) and 33,507 (5 candidates) Da, respectively. The expression of selected 13 candidates (0.2 and 1.0 % error tolerances) were analyzed using real time RT-PCR. The proteins that differentially regulated during oocyte in vitro maturation in the pigs may be potential biomarkers of oocyte maturation and quality.

• 한국수정란이식학회지
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• 제9권1호
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• pp.85-93
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• 1994

This study was carried out to investigate the effects of gonadotropins added during maturation of porcine oocytes on the in vitro maturation(IVM), in vitro fertilization(IVF) and developmental potential of embryos. The follicular oocytes were cultured in TCM-199 medium containing different combination of gonadotropins(5$\mu$g /ml FSR or 1OIU /ml PMSG and 1O$\mu$g /ml LH or 1OIU /ml hCG), 10% FCS and 10% PFF for 36~48h in a incubator with 5% $CO_2$ in Air at 39$^{\circ}C$ and then matured oocytes were again cultured to 120h after IVF for 6~7h with heparin(100$\mu$g /m')-treated sperm. When the oocytes were matured for 42brs in the medium containing FSH+LH, FSH+hCG, PMSG+LH or PMSG+hCG, the JVF rate of each treatment was 50.0%, 52.9%, 66.7% and 70.0%, respectively. The highest CEI (cumulus cell expansion index) was obtained from PMSG+hCG-added medium and the highest polyspermic penetration resulted from FSH+LH-added medium. The cleavage of IVF oocytes derived from hormone added IVM was significantly(P<0.05) promoted by PMSG+hCG and the cleavage rate after 36-h, 42-h and 48-h maturation aws 53.0%, 56.7% and 45.6%, respectively. The highest developmental potential resulted from the oocytes derived from PMSG+LH -added IVM.

• Reproductive and Developmental Biology
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• 제29권2호
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• pp.121-125
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• 2005

The present study was performed to determine the ability of canine oocytes to achieve nuclear maturation according to oocyte diameter and different culture environments. All of the collected oocytes were classified by grade 1 to 3 and by their diameters such as $<100{\mu}m,\;<100{\mu}m\;to\;<110{\mu}m,\;<110{\mu}m,\;to\;<120{\mu}m,\;>120{\mu}m,$. Oocytes were cultured in culture medium supplemented with $10\%\;FBS,\;0.4\%\;BSA,\;10\%$ porcine follicular fluid (pFF), $10\%$ canine serum (CS), or $10\%$ canine estrus serum (CES). The mean number of oocytes recovered from estrus status ovaries was significantly higher than that of anestrus status ovaries (p<0.01). The maturation rate of grade 1 oocytes $(>120{\mu}m)$ was significantly higher than that of the other groups (p<0.05). Nuclear maturation to MI to MII in diameter of $>110{\mu}m$ groups was significantly higher than that in $<100{\mu}m$ group (p<0.05). The oocytes cultured in $10\%$ FBS­supplemented group were significantly higher rate of GVBD compared to the other supplemented groups (p<0.05), and oocytes maturation to MI to MII in $10\%$ FBS-, $0.4\%$ BSA-, and $10\%$ pFF-supplemented groups were significantly higher than those in $10\%$ CS-supplemented group (p<0.05). Based on these results, the estrus status and the size of oocyte affect positively to improve nuclear maturation of canine immature oocytes in vitro. Among several protein sources, porcine follicular fluid was the most effective supplementation to culture medium to achieve higher in vitro maturation rate.

• 한국가축번식학회지
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• 제15권3호
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• pp.221-224
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• 1991

This study was undertaken to evaluate the effect of rabbit peritioneal fluid(rPF) on in vitro maturtion of porcine follicular oocytes. From does 20h after hCG injection, rPF was aspirated aseptically at laparatomy, and then centrifuged, filtrated, and preincubated immediately for 12h. Porcine follicular oocytes isolated from ovaries of slaughtered animals were incubated in TCM-HEPES＋10% FCS, TCM-HEPES＋rPF(v/v, 50/50), or rPE only and examined the nuclear maturation after aceto-orcein or hochest staining. After identifying the optimal incubation time, this experiment was repeated for 5 times. Under the TCM-HEPES containing hormones and serum codition, the time range of porcine follicular oocyte maturation was 38 to 44 hours and the optimal time of maturation of follicular oocyte in vitro was 42 hour cultivation, respectively. The maturatin rates(89.4% and 92.7%) of porcine follicular oocytes cultured in the media with 50% rPF or only rPF were signifciantly higher thanthat (84.6%) of oocytes cultured with TCM-HEPES, respectively. These results suggest that the unknown components(s) of rPF promoted in vitro maturation of porcine follicular oocytes.

• 한국동물생명공학회지
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• 제34권3호
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• pp.205-211
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• 2019

In this study, we investigated the possibility of using mouse embryonic stem cell conditioned medium (ESCM) and embryonic stem cell medium (ESM) for in vitro maturation in the efficient in vitro production of blastocysts from porcine follicular oocyte. Depending on the concentration of supplement of ESCM added to the NCSU-23 solution did not affect 2-cell development rates and blastocysts development. However, in particular, the survival rate (10 days of culture) of blastocyst was significantly higher than that of the control group as the additive concentration (30%) increased (p < 0.05). The survival rate of blastocysts showed a similar tendency even with addition of ESM (30%) alone. On the other hand, the duration of the addition of these additives during IVM (0-44 h) was that the IVM I period (0-22 h) were more effective than the IVM II period (22-44 h). Thus, the effect of these additives is probably due to the combination of the various physiologically active substances of ESCM or the appropriate amino acids and vitamins of ESM. In particular, these additives were more effective during the first half (IVM I) of in vitro maturation. In summary, optimization of ESCM or ESM supplementation may improve in vitro maturation of porcine oocyte and affect developmental competency. Therefore, if more efficient methods of adding ESCM or ESM to basal culture medium can be developed during in vitro maturation of porcine follicle oocytes, high quality blastocysts will be developed from low porcine follicular oocyte compared to other domestic animals.

• Clinical and Experimental Reproductive Medicine
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• 제49권2호
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• pp.149-158
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• 2022

Objective: Optimizing culture media for the incubation of immature oocytes is a vital strategy to increase the oocyte maturation rate during in vitro maturation (IVM) programs. This study evaluated the IVM and fertilization rates of human germinal vesicle (GV) and metaphase I (MI) oocytes using two different maturation media (commercial and homemade) with or without growth differentiation factor 9-β (GDF9-β). supplementation. Methods: Immature oocytes from intracytoplasmic sperm injection (ICSI) cycles were collected and assigned to one of two IVM culture media (commercial or homemade; cleavage-stage base). After maturation, MII oocytes were examined under an inverted microscope for the presence of the polar body, zona pellucida (ZP) birefringence, and meiotic spindle (MS) visualization after maturation in four conditions (commercial or homemade medium, with or without GDF9-β. ICSI was done for matured oocytes, and fertilization was confirmed by the visualization of two distinct pronuclei and two polar bodies. Results: No significant differences were found between the two culture media in terms of the time and rate of oocyte maturation or the rate of fertilization (p>0.05). Growth factor supplementation increased the 24-hour maturation rate for both GV and MI oocytes only in homemade medium. The maturation rate after 24 hours was higher for MI oocytes (p<0.05). Similar results were observed for MS visualization and ZP structure in both types of media (p>0.05). Conclusion: Higher rates of oocyte maturation and fertilization were observed after application of homemade medium supplemented with GDF9-β. Therefore, this combination may be recommended as an alternative for clinical IVM programs.

• 한국수정란이식학회지
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• 제9권3호
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• pp.261-268
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• 1994

Immatured bovine follicular oocytes added with serum, hormones, granulosa cells and bovine oviduct epithelium cells were fertilized in vitro after in vitro maturation. In vitro maturation and early development capacity were examined and IVF-derived embryos were transferred and to recipients and effects of sperm treatment on in vitro capacitation were investigated. The rate of in vitro maturation was improved when they were co-culutred with granulosa cells in the TCM199 medium added with 10% FCS and hormones. The percentage of acrosome reaction was not differed between sperm treatments and sperm of above 25% under-went AR during 30 min preincubation with caffeine and heparin. The cleavage rate of oocytes in vitro fertilized in TCM199 medium added with 10% FCS and hormones, GC or BOEG higher than that in medium with 10% FCS and GC. But the rate was not significantly different between GC and BOEG The cleavage of rate oocytes cultured in medium containing serum, hormones and BOEG was 80.2% and more embryos were developed to Blastocyst (17.3%). The selected embryos were transferred to 9 recipients by surgical or nonsurgical method but did not result in pregnancy.

• Journal of Magnetics
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• 제19권3호
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• pp.241-247
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• 2014

The objective of this study was to study the effect of magnetized water on porcine cumulus cell-oocyte complexes (COCs). Oocytes obtained from female pig were cultured in a medium magnetized at 0, 2000, 4000, and 6000 Gauss (G) for 5 minutes using the neodymium magnet. Subsequently, intracellular hydrogen peroxide ($H_2O_2$) concentration, glutathione (GSH) activity, oocyte membrane integrity, anti-apoptosis factor Bcl-xL expression, and nuclear maturation were analyzed. The intracellular $H_2O_2$ levels in COCs cultured for 44 hours were not significantly different among the variously magnetized samples. However, GSH activity were significantly higher in the magnetized samples compared to the 0 G sample. The Bcl-xL mRNA expression in COCs cultured for 44 hours was higher in the 4000 G sample than other treatment groups. Membrane damage in COCs cultured for 22 and 44 hours was significantly lower in 4000 G group than control group. On the other hand, nuclear stages as maturation indicator significantly increased in 2000, 4000, and 6000 G groups compared to 0 G group. These results indicate that incubation of porcine oocytes and cumulus cells in magnetized medium improves intracellular GSH levels, membrane integrity and nuclear maturation, and inhibits apoptosis in vitro.

• Clinical and Experimental Reproductive Medicine
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• 제20권2호
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• pp.157-160
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• 1993

The human follicular f1uids(F.F.) may be considered to contribute the maturation of the oocytes on the in vitro culture. To investigate the effects of epidermal growth factor (E.G.F.), which is present in mature and immature follicular fluids, we had experiments of mouse oocytes maturation in vitro. The endpoints assayed were rated as percentage of oocytes undergoing germinal vesicle breakdown(G.V.B.D.) and polar body(P.B.) formation at 12 hours after in vitro culture. The rates of G.B.B.D. were 87% in mature F.F. 68% in immature F.F. and 78% in Ham's F-10 medium respectively. And overall the mature F.F. seem to stimulate on in vitro oocyte maturation compared with either immature F.F. or Ham's F-10 medium. As the concentration of addition of E.G.F. in immature F.F., the rates of G.V.B.D. and P.B. formation were 82 %, 23% in addition with 2 ng/ml while 84%, 32% in addition with 4 ng/ml respectivly. And at the concentration of addition of E.G.F. in Ham's F-10 media as well, the rates of G.V.B.D. and P.B. formation were 84%, 40% and 82%, 44% in addition with each 2ng, 4ng. AccordinglY there was no influence on the oocytes maturation at the addition of E.G.F. to each immature F.F. and Ham's F-10 medium. In conclusion, the E.G.F. is not able to induce oocyte maturation independent of it's effects in immature F.F. and Ham's F-10 media.