• Journal of Life Science
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• v.16 no.4
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• pp.571-578
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• 2006

HtrA2/Omi, a mitochondrial trypsin-like serine protease, is pivotal in regulating apoptotic cell death. Several lines of recent evidence suggest that HtrA2 is associated with the pathogenesis of neurodegenerative disorders; however, the physiological function of HtrA2 still remains elusive. For studying physiological function of HtrA2 in depth, it is necessary to develop a suitable expression system in the model animal. We therefore utilized the zebrafish as a model animal to establish expression of human HtrA2 (hHtrA2) in vivo. For expression of mature HtrA2 as GFP fusion in zebrafish embryos, the HtrA2 (WT) or (S306A) cDNAs with the C-terminal GFP tag were inserted into the pCS2+ plasmid. Expression patterns of HtrA2 in HEK293 cells were first monitored by immunofluorescence staining and immunoblot assays, showing approximately 64 kDa of the HtrA2-GFP fusion proteins. Subsequently, the hHtrA2 plasmid DNA or in vitro transcribed mRNA was microinjected into zebrafish embryos. The expression patterns of HtrA2 in Zebrafish embryos were monitored by GFP fluorescence in 24 hours-post-fertilization (hpf). Although expression patterns of HtrA2-GFP in developing embryos were different between the injected DNA and mRNA, both nucleic acids revealed good expression levels to further study the physiological role of HtrA2 in vivo. This study provides a suitable condition for expressing hHtrA2 in the zebrafish embryos as well as a method for generating useful system to investigate physiological properties of the specific human genes.

• Korean Journal of Animal Reproduction
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• v.25 no.2
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• pp.131-138
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• 2001

This experiment was designed to evaluate effects of nonspecific immunostimulator(NIS) Barodon-FX(equation omitted), anionic alkali mineral complex and far-infrared radiation solution on in vivo-produced mouse and in vitro-produced bovine embryos to blastocyst development. Proportion of mouse embryos developing into blastocyst was not greater in BSA- and Barodon-added medium than in BSA-control, but there was signifcantly different(P ＜ 0.05) in hatching and hatched blastocyst development between 0.25% Barodon-and PVP-contained medium(54.7%) than PVP-control(32.5%). BOEC and GC resulted in higher proliferation rate(24∼40% and 17∼22%, respectively) in 0.25∼0.5% Barodon-added medium than in controls, but proliferation of GC and CC greatly decreased in 1∼2% Barodon-added medium. Effect of Barodon on cell proliferation greatly varied among somatic cells. Proportion of early bovine embryos developing into morula and blastocyst was significantly greater(P ＜ 0.05) in 0.5% Barodon-added medium(50% and 63.6%) than in control(31.6% and 27.4%) under co-culture with BOEC and GC, but developmental rate was not different between other Barodon treatments and control. These data indicate that effect of Barodon on cell proliferation significantly varied between somatic cells and that addition of 0.5% Barodon in BOEC-coculture system may further improve blastocyst development in early bovine embryos.

• Korean Journal of Veterinary Research
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• v.33 no.3
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• pp.547-554
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• 1993

This study was carried out to investigate the best condition for in vitro and in vivo culture after freezing and thawing of nuclear transplant 2 cell embryos. When nuclear transplant embryos were submitted to electrofusion, the significantly higher fusion rates of 2 cell donor nuclei were achieved at the electric field strength of DC 1.5 kV/cm for 100 and $150{\mu}sec$, DC 2.0 kV/cm for 100 and $150{\mu}sec$ than DC 1.0 kV/cm for 100 and $150{\mu}sec$(p<0.01). The significantly higher fusion rates of 4 cell donor nuclei were achievecl at DC 2.0 kV/cm for 100 and $150{\mu}sec$ than DC 1.0 kV/cm for 100 and $150{\mu}sec$(p<0.01). The fusion rates in 8 cell donor nuclei were 94.2~99.3%. The developmental potency to blastocyst in 2 cell donor nuclei was significantly higher in DC 2.0 kV/cm for $150{\mu}sec$ treated group(p<0.01). The significantly higher developmental potency to blastocyst in 4 cell donor nuclei were achieved at the electric field strength of DC 2.0 kV/cm for $150{\mu}sec$ than DC 1.5 kV/cm for 100 and $150{\mu}sec$, DC 2.0 kV/cm for $100{\mu}sec$ treated group(p<0.01). The develop mental potency to blastocyst in 8 cell donor nuclei was significantly higher in DC 2.0 kV/cm for $100{\mu}sec$ treated group(p<0.01). The developmental potency to blastocyst after nuclear transplantation was significantly higher in 2 cell donor nuclei than in 8 cell donor nuclei(p<0.01). When the recovered embryos in normal morphology were cultured in vitro, there were no significant differences in the developmental potency to blastocyst between the freezing methods and the concentrations of cryoprotectant(p<0.01). The production rates of offspring after transfer of nuclear transplant embryos to recipient mouse were no significant difference in 2, 4 and 8 cell donor nuclei.

• Korean Journal of Animal Reproduction
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• v.22 no.2
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• pp.171-176
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• 1998

This study was carried out to confirm whether the in vivo developmental potential of mouse hatched blastocysts (HBs) can be obtained by vitrification method using the cryoprotectant EFS35. The HBs ($\theta$ 130$\mu\textrm{m}$) were cultured in vitro until day 5 from zygotes produced in vivo and were equilibrated in 10% ethylene glycol(EG) for 5 min, and then were exposed or vitrified in EFS35 (35% EG, 18% Ficoll and 0.3M sucrose). After 30 min thawing, re-expanding HBs were transferred into one or both uterine horns of pseudopregnant recipients on day 3 (4~6 embryos /horn). Pregnancy rates of recipients and implantation were assessed by autopsy on day 15 of gestation. The results obtained in these experiments were summarized as follows : After thaw-ing, in vitro survival of HBs was not significantly different between exposed (65.5%) and vitrified(54.5%) group. Also, when the in vivo development potential was examined, total implantation was not different between control (58.5% ) and vitrified (41.0%) group, although the live fetus formation of vitrified group (24.0%) was significantly lower than that of control (58.3%) group (p< 0.05). These results suggested that vitrification freezing method of mouse HBs using EFS35 can be used to make wide the utility of embryo transfer of the more embryos produced in vitro.

• Journal of Microbiology and Biotechnology
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• v.11 no.4
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• pp.656-662
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• 2001

Angiogenesis is known as a crucial process in the growth and spreading of tumor cells. Accordingly, the effective inhibition of this process would appear to be a promising way to cure angiogenesis-related diseases, including cancer. This study demonstrates that acalycixenolide E (AX-E) from the marine organism Acalycigorgia inermis exhibits a potent anti-angiogenic activity both in vitro and in vivo. AX-E inhibits the bFGF-induced proliferation of HUVECs in a dose dependent manner, along with the bFGF-induced migration, invasion, and tube formation of HUVECs. Moreover, AX-E potently inhibits the in vivo neovascularization of the chorioallantoic membranes (CAMs) of growing chick embryos. interestingly, AX-E suppresses the expression of metalloproteases 2 and 9, yet shows no effect on their activities. The novel chemical structure and potent anti-angiogenic activity of AX-E will be of great value in elucidating the molecular mechanism of angiogenesis as well as in the development of a novel anti-angiogenic drug.

• Journal of Embryo Transfer
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• v.21 no.3
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• pp.225-231
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• 2006

The objective of this study was to determine changes in serum hormone concentrations, blood chemical values and recovery rate of in vivo embryos during the estrous cycle following super-ovulation treatments in Jeju black cows. Superovulation was induced by subcutaneous administration of FSH twice a day for 4 days. Serum hormones were assayed by radioimmunoassay (RIA) and blood chemical values were analyzed by blood analyser system. Embryos were collected from all treated black cows using nonsurgical technique on day 7 after artificial insemination (AI). The results of this study were summarized as follows: 1. The progesterone concentrations were $7.2{\pm}3.8ng/ml$ at day -11 and $0.3{\pm}0.1ng/mL$ at day 0 (Day 0 is the first day of AI). The estradiol concentrations were $10.6{\pm}4.48pg/ml$ at day -11 and $15.0{\pm}2.2pg/ml$ at day 0. The lowest level of progesterone was measured at day 0. The highest levels of estradiol was measured at day 0. 2. The blood chemical values of treated black cows were no significant differences in normal cow values. 3. Sixty two embryos were collected in 12 black cows. Among the collected embryos, 37 embryos (59.7%) could be transferred into recipients. These results would be used as the basic informations for changing patterns of hormonal level and blood biochemistry in Jeju black cow with superovulation.

• Fisheries and Aquatic Sciences
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• v.25 no.2
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• pp.90-100
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• 2022

Cardiovascular diseases (CVDs) have posed serious public health problems, accounting for nearly 30% of mortality worldwide and their incidence is still increasing. Therefore, new treatment resources are necessary to prevent or manage the ever-increasing population of patients with CVDs. Sea cucumber is well known for its medical and health benefit effects, but it is not well known what/how effect it has on vascular disease. In the present study, we examined the protect effect of sea cucumber, Apostichopus japonicus 80% ethanol extract (AJE) on zebrafish embryo with the stimulation of free fatty acid, palmitate (PA). In vivo study showed that AJE can attenuate PA-induced toxicity through relieving the rapid heartbeat, increasing the survival rate and reducing the malformation in both wild type and Tg (fli1a:eGFP) transgenic zebrafish lines. Additionally, compare with PA treated embryos, the yolk sac area, body length, axial vascular segment (AVS) and intersegmental vessel (ISV) of the co-treatment group of AJE and PA were comparable to the control group. Moreover, AJE lowered the expression of inducible nitric oxide synthase (iNOS), nitric oxide (NO) and inflammation-related genes induced by PA, and inhibited PA-induced vascular development disorders. Our data preliminarily verify that AJE could be a candidate resource for the prevention or therapy of CVDs.

• Journal of Embryo Transfer
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• v.25 no.4
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• pp.221-227
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• 2010

Early cleavage is a reliable prognostic tool for successful embryo transfer in assisted reproduction because early cleaved embryo show better pregnancy rate after transfer. There for, preparation of good embryo recipient is important factor to optimize efficiency of pig cloning. The present study was performed to evaluate the effect of early cleavage on the in vivo development of cloned embryos and to analyze breed, parity and estrous synchrony to optimize recipient for pig cloning. In vitro matured porcine oocytes derived from local slaughterhouse and fibroblasts derived from miniature pig fetuses were used for somatic cell nuclear transfer (SCNT). Reconstructed embryos were transferred to recipient pigs on the same day of SCNT or after 1~2 days of in vitro culture for selecting early cleaved embryos. Breed, parity and date of standing estrous of recipients were recorded for analysis. After 25~35 days after embryo transfer pregnancy was diagnosed using ultrasonography, and pregnant recipients were monitored till delivery. Between purebred and crossbred, no significant difference was founded in both pregnancy and delivery rates. However, early cleaved embryos showed significantly higher pregnancy (46.2%) and delivery (12.8%) rates compared to non-selectively transferred group (24.8% and 4.5%, respectively). The results also showed that the recipients showing standing estrous on the same day of SCNT and less than 4 parities were most suitable for pig cloning.

• Asian-Australasian Journal of Animal Sciences
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• v.4 no.1
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• pp.73-78
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• 1991

Sixty-one 5-11 month-old California, Chinchilla and New Zealand White rabbit were employed in this investigation. Thirty-three does were superovulated by injecting FSH/HCG subcutaneously or intravenously and then sacrificed at different hours after mating. The ova were collected from the fallopian tubes with Ham's F-10 medium supplemented with 0.4% bovine serum albumin (BSA) and 1% pregnant rabbit serum (PRS). Embryos were examined under an inverted DIC microscopy for observing the stage of development. We have found that the fertilized ova formed pronuclei at 19 - 20 hr postcoitus. Approximately at 26, 64 - 78 and 84 - 88 hr after mating, the fertilized ova cleaved further to 2-cell, morulae and blastocyst stage respectively. Another 28 does were allocated to the gene transfer study. Fourteen of the 28 does were sacrificed at 19 - 20 hr to donate the pronuclear stage ova for gene injection. The other 14 does were induced to pseudopreganacy by injection of 100 IU HCG intravenous as recipients. Four hundreds and seventeen ova were injected totally and 212 gene injected zygotes were transferred into the recipient oviducts. Five recipients became pregnant and 10 fetuses were obtained. Eight of the 10 fetuses were analysed for gene incorporation, but none of them were transgenic.

• Journal of Veterinary Clinics
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• v.27 no.2
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• pp.154-158
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• 2010

Embryo Transfer (ET) is one of the assisted reproductive technologies and a useful tool for improving herds. The purpose of this study is to produce the calves using frozen embryos which were produced in the top one percent Holstein in Canada by ET. One hundred seventeen recipients were used for surrogate mothers and seventy cows were diagnosed to be pregnant. Fifty seven calves were born successfully and thirteen out of them failed to produce viable calves (abortion: 4, stillbirth: 9). Their gestational length, birth body weight and sex ratio for all the viable calves(n = 57) were $278.1{\pm}3.6$ days (range: from 271 to 286 days), $44.0{\pm}3.0\;kg$ (range: from 37 to 49 kg) and 57.9 vs. 42.1 % (male 33 and female 24), respectively. Microsatellite analysis confirmed that they were derived from frozen embryos. In conclusion, this study demonstrated that viable calves derived from frozen-thawed embryos from Canada were born by ET.