• Journal of Embryo Transfer
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• v.5 no.2
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• pp.11-22
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• 1990

Remarkable progress has recently been made in embryo transfer technology, resulting in the birth of IVF and nuclear transfer offsprings in swine. However, further progress of the technology to (I) make a safe, effective and economic estrual-cycle synchronization compound, (2) regulate each step of sperm capacitation (3) induce monospermic fertilization, (4) in vitro grow and mature oocytes, (5) fertilize the oocytes efficently, (6) culture the oocytes to the blastocyst stage in defined media, (7) produce multiply copies of embryos with superior genetic merit, (8) preselect the sex of these superior offsprings, and (9) preserve embryos by freezing and storage in liquid nitrogen is required before this promising technology is applied routinely to swine for practical use.

• Korean Journal of Animal Reproduction
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• v.14 no.1
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• pp.73-83
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• 1990

These studies were carried out to find the proper conditions for in vitro maturation and fertilization of bovine follicular oocytes and culture methods capable of further developing early embryos. For these objectives, the cleavage rate of oocytes matured and fertilized in vitro was investigated under medium supplemented with hormones and estrous cow serum and season of oocytes collection as well as different cumulus cell stage before insemination. Finally, 2~8 cell embryos were cultured in in vitro and in vitro culture system to investigate developmental capacity into morula. 1. Cleavage rate of oocytes matured in vitro was 27%(20/73) for A(LH＋FSH＋estradil-17$\beta$＋10% FBS), 38%(27/71) for B(LH＋10% ECS) and 27%(15/56) for C(10% ECS), respectively. Supplement B showed more higher rate and 4~8 cell embryos were also obtained much more in this group(67%, 18/27). In vitro maturation rate of follicular oocytes cultured in TCM 199 supplemented withLH and 10% ECS was 88%(75/85). 2. Cleavage rate(15%, 10/65) of oocytes collected in summer was significantly lower than in fall(47%, 42/89). 3. Cleavage rate(15%, 10/65) of oocytes collected in summer was significantly lower than in fall(47%, 42/89). 3. Cleavage rate(15%, 10/65) of oocytes with partially removed cumulus cell before insemination was more higher than that(44%, 27/62) of oocytes with intact cumulus cell. 4. The frequency of development from early cleaved embryos into morula was 6%(4/65), 12%(4/33) for co-culture of cumulus cell monolayer and bovine oviduct epithelial cell monolayer, respectively and 25%(6/25) in ligated rabbit oviduct.

• Korean Journal of Animal Reproduction
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• v.19 no.4
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• pp.259-264
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• 1996

This study was carried out to investigate on the survival rates and in vitro developmental rates of bisected bovine embryos were by manipulator and micropipette. Bisected embryos were co-cultured in 20% FCS(v/v)＋TCM-199 media containing hormones, oviductal epithelial cells and cumulus cells 0 to 72 hrs after bisection. Survival rate and in vitro fertilization rate were defined as development rate on in vitro culture or FDA-test. The results are summarized as follows ; 1. The in vitro developmental rate of biseciton embryos co-cultured in 20% FCS＋TCM-199 medium containing PMSG, hCG, PMSG＋hCG, hCG＋$\beta$-estradiol 0 to 20 hrs and 20 to 40 hrs were 36.7, 26.7, 33.3, 40.0, and 30.0, and 30.0, 33.3, 30.0, 26.7, and 26.7%, respectively. The survival rate of bisection embryos co-cultured in TCM-199 medium containing hormones was significantly higher than that of non co-culture(25.0%). 2. The in vitro developmental rates of bisection embryos co-cultured in 20% FCS＋TCM-199 medium containing oviductal epithelial cells 4 to 5 hrs and 20 to 24 hrs were 40.0 and 33.3%, respectively. The survival rate of bisection embryos co-cultured in TCM-199 medium containing oviductal epithelial cells was significantly higher than that of non co-culture(25.0%). 3. The in vitro developmental rates of bisection embryos co-cultured in 20% FCS＋TCM-199 medium containing cumulus cells 4 to 5 hrs and 20 to 24 hrs were 43.3 and 36.7%, respectively. The survival rate of bisection embryos co-cultured in TCM-199 medium containing cumulus cells was significantly higher than that of non co-culture (25.0%).

• Reproductive and Developmental Biology
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• v.39 no.3
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• pp.49-57
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• 2015

In order to achieve successful in vitro production of embryo, it is necessary to establish intrauterine environment during in vitro culture. Thus, this study was investigated to establish embryo culture system using co-incubated collagen matrix gel (CM) with endometrial epithelial cells (EC). Endometrial epithelial cells were isolated from porcine endometrium at follicular phase, the cells seeded in insert dish for co-incubation with CM-coated culture dish. Then, culture media treated with/without 2.0 IU/ml hCG or 10 ng/ml $IL-1{\beta}$. After incubation for 24 h, the co-incubated insert dishes were removed from CM-coated culture dish before embryo culture. Embryos at 48 h after in vitro fertilization (IVF) were cultured on the dish for 120 h with porcine zygote medium. We determined PTGS-2 expression in the ECs, VEGF protein in co-incubated CM with EC and observed cleavage rate and blastocyst development of embryos at 168 h after IVF. In result, expression of PTGS-2 was higher at co-incubated EC with hCG and $IL-1{\beta}$ groups than EC without hCG and $IL-1{\beta}$. The VEGF protein was detected at co-incubated CM with EC, EC treated with hCG and $IL-1{\beta}$ groups higher than CM group. Also, cleavage rate was no significantly difference among all group, however, blastocyst development was significantly higher in co-incubated CM with EC treated with hCG group than un-treated groups (p<0.05). Therefore, we suggest that novel embryo culture system using co-incubated collagen matrix gel with endometrial epithelial cells treated with $IL-1{\beta}$ is beneficial and useful for enhancing the production of porcine blastocysts in vitro.

• Journal of The Korean Society of Grassland and Forage Science
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• v.33 no.2
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• pp.105-110
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• 2013

This study was carried out to investigate the effect of nitrogen (N) fertilization on the forage growth, production and quality of native reed (Phragmites communis) grasses. Field experiments were conducted in Cheonan and in Ansan, 2012. Treatments were control (no N fertilization), 50 kg/ha and 100 kg/ha in Cheonan plots (fertilization on April $30^{th}$, and harvest on June $21^{st}$). Treatments in Ansan plots were control (no N fertilization) and 60 kg/ha (fertilization on May first, and harvest on August first). Forage growth and leaf colors were improved in N fertilized plots. However, the drymatter (DM) percentage was slightly decreased with N fertilization. Forage yields, in terms of DM, crude protein (CP) and digestible DM (DDM), were significantly increased with N fertilization in both sites. In Cheonan, DM, CP and DDM yields per ha were 4,026 kg, 235 kg and 1,850 kg, respectively, in the control plot, and were 4,658 kg, 306 kg and 2,388 kg, respectively, in the N 50 kg plot, and 5,622 kg, 446 kg and 3,143 kg, respectively, in the N 100 kg plot. In Ansan, DM, CP and DDM yields per ha were 2,802 kg, 177 kg and 1,288 kg, respectively, in the control plot, and were 3,876 kg, 294 kg and 1,853 kg, respectively, in the N 60 kg plot. Forage quality in terms of CP content, in vitro DM digestibility (IVDMD) and relative feed value (RFV) were also increased with N fertilization in both sites. In Cheonan, the CP content, IVDMD and RFV were 5.85%, 45.96% and 64.5 (grade 5), respectively, in the control plot, 6.58%, 51.27% and 72.3 (grade 5), respectively, in the N 50 kg plot, and 7.94%, 55.91% and 72.7 (grade 5), respectively, in the N 100 kg plot. In Ansan, the CP content, IVDMD and RFV were 6.30%, 45.98% and 70.2 (grade 5), respectively, in the control plot, and 7.59%, 47.80% and 78.3 (grade 4), respectively, in the N 60 kg plot. In conclusion, N fertilization of 60~100 kg/ha was desirable for greater forage production, with a higher quality of native Phragmites communis achievable. This should only be applied if the fertilization area is not located at a riverside/streamside or in riparian land where there is a high risk of water pollution by fertilization.

• Journal of Embryo Transfer
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• v.9 no.1
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• pp.15-21
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• 1994

This experiment was investigated the effect of cell stage of embryos at 48 hours post-insemination On in vitro development of IVF embryos. The ovaries of Korean native cows or heifers were obtained from an abattoir and kept on 25 to 28$^{\circ}C$ and transported to laboratorty within 2 hrs. The oocytes were matured in vitro(IVM) for 24 hrs. in TGM-199 supplemented with 35 $\mu$g/$m\ell$ FSH, 10 $\mu$g/$m\ell$ LH, 1 $\mu$g/$m\ell$ estradiol-17$\beta$ and granulosa cells at 39$^{\circ}C$ under 5% $CO_2$ in air. They were fertilized in vitro(IVF) by epididymal spermatozoa treated with heparin for 24 hrs. , and then the zygotes were co-cultured in vitro(IVC) with bovine oviductal epithelial cells for 7 to 9 days. At 48 hrs. post-insemination, the embryos were classfied into 5 to 8-cell, 3 to 4-cell or 2-cell stage and then were co-cultured in vitro(IVC) with bovine oviductal epithelial cells until the embyos reached blastocyst stage. Embryos developed to blastocyst stage were stained with Hoechst 33342 for cell counting. The embryos of 5 to 8-cell stage at 48 hrs. post-insemination with grade I oocytes were significantly (P<0.05) better developed to blastocysts(63.0%) than 3 to 4-cell(42.0%) and 2-cell stage(2.7%) embryos which delayed in the early cleavage, and those embryos cleaved faster in the very early stage seemed to develop to blastocysts earlier. These results indicate that the embryos cleaved faster at 48 hrs. post-insemination seemed to develop to blastocysts earlier.

• Development and Reproduction
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• v.6 no.1
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• pp.17-23
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• 2002

In oocyte retrieval, a vein anesthetic drug is commonly used for induction and maintenance of general anesthesia. Propofol and Thiopental sodium are frequently used for ultrasound-guided transvaginal oocyte retrieval. The present study aimed to assess the effects of Propofol and Thiopental on in vitro fertilization(IVF). Immature porcine oocytes were exposed to various concentrations ot Propofo1 and Thiopental sodium. The rates of oocyte maturation, fertilization and development were observed. The parthenogenetic effects of the anesthetics were also evaluated. The rate of oocyte maturation after exposure to high concentrations of the anesthetics for long time was significantly higher than that of the control. But the rate of fertilization after long-time exposure to the high concentration of the anesthetic drugs was significantly lower than that of the control. The results support that Propofo1 serves like other anesthetics described, as a parthenogenetic activator. Oocytes exposed to Thiopental sodium showed decreased rates of maturation and fertilization. These results suggest that usage of optimum concentration of anesthetic drug is important in increasing the rates of oocyte maturation, fertilization and development in IVF.

• 대한생식의학회:학술대회논문집
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• 2006.11a
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• pp.139.1-139.1
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• 2006

• Proceedings of the KSAR Conference
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• 2001.10a
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• pp.44-46
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• 2001

In vitro produced (IVP) embryos produced by in vitro fertilization (IVF) often exhibit wide variations in developmental competence and viability, considerably more than are exhibited by embryos that develop in vivo. These anomalies in IVP embryos may be due to heterogeneity of oocyte quality, suboptimal culture conditions, disturbances in gene expression, or most likely a combination of these factors (Ho et al., 1994; Roth et al., 1994; McKiernan and Bavister, 1998; Hasler, 1998; Schramm and Bavister, 1999; Doherty et al., 2000; Hyttel et al., 2000; Niemann and Wrenzycki, 2000; Wrenzycki et al., 2001). In research studies or in clinical applications with domesticated animals, cats, non-human primates and humans, oocytes used for IVF are usually collected from a heterogeneous cohort of ovarian follicles that include oocytes which normally might not be ovulated and/or are deficient in developmental competence. (omitted)

• Korean Journal of Animal Reproduction
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• v.17 no.3
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• pp.249-255
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• 1993

Canine follicular oocytes were used to establish a reliable system for maturation and fertilization in vitro. Ovaries were obtained from either slaughter house or hormone-primed bitches of mixed breeds. The oocytes were recovered by mincing the ovaries in M2＋BSA. Good quality of oocyte-cumulus complexes (OCCs) were selected and cultured in TCM 199 containing 15% fetal calf serum(FCS) for 24~56 h in an atmosphere of 5% CO2 at 39$^{\circ}C$. Maturation rate of follicular oocytes was >87% showing metaphase I. Unlike other domestic animals the cumulus expansion did not occur fully in canine OCCs although minimum expansion was found between the cumulus cells and corona radiata cells, the clear nuclear morphology was presented for the first time by rapid staining. The IVM system used in this study may be useful to obtain fully maturated metaphase I oocyte in dog.