• Journal of Embryo Transfer
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• v.30 no.1
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• pp.23-31
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• 2015

Catalpol, an iridoid glucoside, isolated from the root of Rehmannia glutinosa Libosch. It possesses a broad range of biological and pharmacological activity including anti-tumor, anti-inflammation and anti-oxidant by acting as a free radical scavenger. Therefore, in this study, the effects of catalpol on blastocyst development, expression levels of reactive oxygen species (ROS) and apoptotic index were investigated in porcine embryos. After in vitro maturation and fertilization, porcine embryos were cultured for 6 days in porcine zygote medium 3 (PZM-3) supplemented with catalpol (0, 100, 200 and $400{\mu}M$, respectively). Blastocyst development not significantly improved in the catalpol treated group when compared with control group. Otherwise, the intracelluar levels of ROS were decreased and the numbers of apoptotic nuclei were reduced in the catalpol ($100{\mu}M$) treated porcine blastocysts (P<0.05). On the other hand, blastocyst development was significantly improved in the catalpol ($100{\mu}M$) treated group when compared with the untreated catalpol group under $H_2O_2$ ($200{\mu}M$) induced oxidative stress (P<0.05). Otherwise, the intracellular levels of ROS in catalpol ($100{\mu}M$) treated group were significantly decreased in the untreated catalpol group under $H_2O_2$ ($200{\mu}M$) induced oxidative stress (P<0.05). Furthermore, the total cell numbers of blastocysts were significantly increased (P<0.05) in the catalpol ($100{\mu}M$) treated group under $H_2O_2$ ($200{\mu}M$) induced oxidative stress, whereas numbers of apoptoic nuclei were significantly reduced (P<0.05). In conclusion, our results indicate that treatment of catalpol may have important implications for improving developmental competence and preimplantation quality of porcine embryos through its anti-oxidant and anti-apoptotic effect.

• Development and Reproduction
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• v.1 no.1
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• pp.79-89
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• 1997

Recent studies have demonstrated that apoptotic cell death plays an important role in the mechanism underlying follicular atresia and luteolysis. However, the mechanisms responsible for initiating these processes have not been elucidated. In in vitro fertilization (IVF) programs, it is highly possible that continuous and repeated administration of FSH/hMG and GnRH agonists for the usage of ovarian hyperstimulation may induce apoptotic death of granulosa cells leading to atresia in the human ovarian follicles. The present study was performed to investigate whether FSH/hMG and GnRh agonists used for a longer period in controlled ovarian hyperstimulation has any effect on the apoptosis of granulosa-luteal (GL) cells obtained from hyperstimulated ovaries. To examine apoptotic cell death in the GL cells, cells were stained with acridie orange followed by observed in some of GL cells. Similar but distinct staining of apoptotic GL cells was observed when the cells were examined by using in situ TUNEL method. The healthy-looking cells with normal nuclear morphology were not stained, whereas cells with pyknotic nuclei or with apoptotic nuclei were intensively stained. After examining the ultrastructural features of GL cells by TEM, it was confirmed that the majority of cells seemed to have normal nuclei while GL cells undergoing apoptotic cel death were rarely found. The DNA extracted from GL cells showed a typical pattern of fragmentation following DNA electrophoretic analysis. We have confirmed that the apoptosis occurs in granulosa-luteal cells obtained from hyperstimulated ovaries. Technically, in situ apoptosis detection method is simple and reproducible and is well suited to identify the quality of oocytes retrieved from hyperstimulated ovaries.

• Reproductive and Developmental Biology
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• v.32 no.2
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• pp.81-87
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• 2008

Production of transgenic animals for studying specific gene has been limited due to a low efficiency, lack of skilled researchers and the need for expensive equipment. Currently, the boar spermatozoa as a vector to deliver exogenous DNA into the oocyte were used to improve the efficiency of transfection rate. In this study, we revealed that the optimal conditions for DNA uptake in spermatozoa by liposome were to 90 min of incubation, $17^{\circ}C$, $10^5$ spermatozoa, 4 ng/ml of exogenous DNA and 0.5% (v/v) liposome, without damage to fertility. In addition, the developmental rate to the blastocyst stage of embryo in control group was significantly higher than those embryos with exogenous DNA and liposome, whereas there were no significant differences in embryo development between the liposome and type of DNA. The transfection rates of embryo using treated spermatozoa with both liposome and circular DNA were higher than those using linear DNA. These findings raise the possibility thattreated spermatozoa with liposome/DNA complexes could be used in in vitro fertilization, and the exogenous DNA transferred into the oocytes. Taken together, we demonstrated that liposome a vector for the uptake of exogenous DNA in boar spermatozoa could improve the efficiency of sperm-mediated gene transfer in creating transgenic pig and the other domestic transgenic animals.

• Clinical and Experimental Reproductive Medicine
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• v.38 no.4
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• pp.234-237
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• 2011

Objective: During stimulated IVF cycles, up to 15% of oocytes are recovered as immature. The purpose of this study was to investigate the trend of oocyte maturity in repeated ovarian stimulation for IVF. Methods: One hundred forty-eight patients were selected who underwent two consecutive IVF cycles using same stimulation protocol during 2008 to 2010. Ovarian stimulation was performed with FSH and human menopausal gonadotropin and flexible GnRH antagonist protocol in both cycles. Oocyte maturity was assessed according to presence of germinal vesicle (GV) and the first polar body. Immature oocyte was defined as GV stage or metaphase I oocyte (GV breakdown with no visible polar body) and cultured up to 48 hours. If matured, they were fertilized with ICSI. Results: Percentages of immature oocytes were 30.8% and 32.9% ($p$=0.466) and IVM rates of immature oocytes were 36.2% and 25.7% ($p$=0.077), respectively. A significant correlation was noted between percentage of immature oocytes in the two cycles (R=0.178, $p$=0.03). Women with >40% immaturity in both cycles (n=21) showed lower fertilization rate of $in$ $vivo$ matured oocytes (56.4% vs. 72.0%, $p$=0.005) and lower pregnancy rate (19.0% vs. 27.1%, $p$=0.454) after the second cycle when compared with women with <40% immaturity (n=70). In both groups, female age, number of total retrieved oocyte and embryos transferred were similar. Conclusion: In repeated ovarian stimulation cycles for IVF, the immature oocyte tended to be retrieved repetitively in consecutive IVF cycles.

• Journal of Embryo Transfer
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• v.28 no.3
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• pp.265-271
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• 2013

Cryopreservation and in vitro fertilization (IVF) protocols are important in genetic studies and applications to transgenic animals. Various studies about boar sperm cryopreservation have been studied for a long time. Those were about the use of extenders, the choice of sugars, the cooling and warming rates. The factors that influence the boar sperm are the dramatic changes in temperatures, osmotic and toxic stresses, and reactive oxygen species (ROS) generation. Among these factors, ROS generation is the main damage to DNA which is a principal genetic material and the most important for the practical applications. So we wondered whether ROS generation could be reduced. In previous study, monothioglycerol (MTG) was essential for the culture of embryo stem cells. Therefore we added MTG in the freezing extender based on lactose-egg yolk (LEY) with trehalose. For the assessment of the frozen-thawed sperm, we focused onmotility, membrane integrity and DNA damage. First, we used a computer-aided sperm analysis system for overall conditions of sperm such as motility and viability. Then we performed the sperm chromatin structure assay for DNA integrity and hypo-osmotic swelling test for membrane integrity. And our result showed the existence of MTG in the freezing extender caused less damage to DNA and higher motility in frozen-thawed boar sperm. Also we checked a relative antioxidant activity of MTG in modified Modena B extender. We concluded that this reagent can activate sperm mitochondria at MTG $0.2{\mu}M$, contribute to sperm motility and DNA integrity but there was no significant difference on membrane integrity. Also antioxidant activity of MTG in modified Modena B extender was proved.

• Korean Journal of Animal Reproduction
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• v.26 no.4
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• pp.329-338
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• 2002

This study was carried out to investigate the developmental rates of embryo reconstructed with different cell type and to estimate correlation of transcriptional level of octamer-binding transcription factor 4 (Oct4) and fibroblast growth factor 4 (FCF4) gene on peri-implantation stage embryos. Donor cells were transferred into perivitelline space of enucleated oocytes. The karyoplast-cytoplast couplets were accom- plished by cell to cell fusion and activated with ionomycin and 6-dimethylaminopurine. Reconstructed embryos were co-cultured with bovine oviduct epithelial cells in CR 1 aa medium. There is no difference in blastocyst formation rate following nuclear transfer UT) with fetal fibroblast cell (16/50; 32.0%), cumulus cell (16/49; 32.6%) and ear cell (17/52; 32.6%). The expression level of Oct4 and FCF4 in peri-implantation bovine embryo derived from in vitro fertilization (IVF) and NT were determined by reverse-transcription polymerase chain reaction (RT-PCR) technique. In peri-implantation of IVF result in a transient increased of FCF4 paralleled by an increased expression of Oct4. However, Oct4 gene was highly expressed in hatching blastocysts derived from NT compared to IVF. Also, FGF4 expression level in hatching blastocysts and outgrowth stage derived from NT was lower than that of IVF. In conclusion, it is suggested that the different transcription patterns observed in nuclear transfer embryos may lead to a lower rate of embryo development, implantation and pregnancy.

• Korean Journal of Organic Agriculture
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• v.6 no.1
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• pp.133-149
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• 1997

Recently, the importance of management and cultivation of grasses has been increased in Korea. Among these cultural practices, the appropriate control of diseases is considered more important than other cultivation techniques such as fertilization and irrigation. The damages of brown patch and large patch caused by Rhizoctonia spp. and Pythium blight caused by Pythium spp. are serious in the major cultivation area of turfgrass in Korea. Since these diseases are difficult to control by agrochemicals, the damages are very serious if these are occured. The periodic spray of agrochemicals, to protect and control these diseases could make some problems of toxicity and environmental pollution as well as rising of non-target diseases. Therefore, the biological methods to control diseases have been required to decrease problems resulted from overuse of agrochemicals, to conserve natural ecosystem, and to control effectively diseases of grasses in the long period. The number of studies about biological control using antagonistic microorganisms have been increased for last half century. However, the application of biological control method has been very limited. In this study, thirteen isolates of R. cerealis, 8 isolates of R. solani and 3 isolates of Phthyn spp. have been isolated from diseased turfgrass in golf course and grass-culture area that have patch and wilting symptoms of zoysia grass and creeping bentgrass. Isolation frequency of R. cerealis and R. solani was high in especially zoysiagrass, while Pythym spp. was isolated from bent grass at low frequency but showed high pathogenicity. Totally, 205 isolates of soil microorganisms were isolated in this study as primary antagonistic microorganism by Herr's triple agar layer plate and dual culture method using rhizosphere of grasses, soil of crop field as the source of antagonistic microorganisms. Among the 205 isolates, 23 isolates were actinomycetes and 182 isolates were bacteria. All of the actinomycetes were isolated by Herr's method. Antagonistic effect of primary isolated microorganisms was tested for in vitro mycelial growth inhibition against pathogenic fungi isolated from grasses and for inhibition of disease occurrence in 24 well tissue culture plate and pot experiment. Then, four isolated of bacteria which are BG23, BG74, BG136 and BG171 were selected as antagonistic microorganisms against soil-born pathogenic fungi of bentgrass.

• Clinical and Experimental Reproductive Medicine
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• v.21 no.3
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• pp.241-245
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• 1994

Despite increasing success rate of IVF, poor response to ovarian stimulation remains a problem. So, attempts to improve ovarian responses, for example, by using combined gonadotropin-releasing hormone analogue(GnRH-a) and human menopausal gonadotropin(hMG) have shown limited success. It is reported that response of granulosa cells in vitro to FSH is stimulated by co-incubation with IGF-l, and IGF-l production can be increased by growth hormone. This suggest that combination regimen of G.H. and hMG may augment follicle recruitment. In fifteen patients who had previous history of poor ovarian response to gonadotropin stimulation after pituitary suppression with mid -luteal GnRH-a, the effectiveness of cotreatment with G.H. in IVF program was evaluated using a combination regimen of G.R. and hMG at Korea University Hospital IVF Clinic. Ovarian responses to gonadotropin stimulation in control and GH-treated cycles assessed by total dose and duration of hMG treatment, follicular development and peak $E_2$ level, number of eggs retrieved, and fertilization rates were also assessed. In each group, serum and follicular fluid IGF-1 concentrations on day of egg collection were measured by RIA after acidification and extraction by reveresed phase chromatography. Patients receiving G.H. required fewer days and ampules of gonadotropins, developed more oocytes, and more embryos transferred. But, the differences were not statistically significant, except the duration of hMG treatment. Our data showed a significantly higher concentration of IGF-l in the serum, not in the follicular fluid, of patients treated with G.H. compared with control group. These data suggest that growth hormone treatment does not improve the ovarian response in women with limited ovarian reserve to gonadotropin stimulation for IVF.

• Journal of East-West Nursing Research
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• v.28 no.2
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• pp.100-108
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• 2022

Korean medicine has traditionally provided a lot of medical care for the treatment of infertility. The combination treatment of Korean medicine and Western medicine can be effective for infertility to improve the problems caused by the extremely low fertility rate and the aging of pregnant women. In relation to female infertility, we reviewed the evidence for Korean medicine treatment for polycystic ovary syndrome, which has recently been increasing interest in female infertility, and the evidence for the collaboration between Korean and Western medicine in the treatment of polycystic ovary syndrome and infertility. Because polycystic ovary syndrome is closely related to metabolic diseases related to endocrine abnormalities, hormone treatment alone has limited therapeutic effects and has side effects. In domestic and international research, herbal medicine and acupuncture in the treatment of infertility have shown good clinical effects based on the hypothesis of various mechanisms of effect, and many cases of successful pregnancy have been reported. Although continuous research through large-scale randomized clinical studies is needed to prove efficacy, the importance of combined treatment of Korean medicine and Western medicine in infertility treatment is increasing in order to solve the extremely low fertility rate. Combining artificial insemination or in vitro fertilization with Korean medicine treatment is highly likely to be effective in increasing pregnancy success and fertility rates. In women with polycystic ovary syndrome, which is recognized as a metabolic disease due to endocrine abnormalities among the causes of female infertility, Korean medicine treatment can increase the pregnancy rate with herbal medicines and acupuncture, etc. Although well-designed large-scale clinical studies are currently lacking and meta-analysis has not provided sufficient evidence, the combination treatment of Korean medicine and Western medicine is more active to increase the pregnancy rate for solving the problem of low fertility.

• Journal of Genetic Medicine
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• v.19 no.1
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• pp.7-13
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• 2022

Purpose: Preimplantation genetic testing for monogenic disorders (PGT-M) has been successfully used to prevent couples with monogenic disorders from passing them on to their child. Charcot-Marie-Tooth Disease (CMT) is a genetic disorder characterized by progressive extremity muscle degeneration and loss of sensory function. For the first time in Korea, we report our experience of applying single nucleotide polymorphism genotyping and karyomapping for PGT-M of CMT disease. Materials and Methods: Prior to clinical PGT-M, preclinical tests were performed using genotypes of affected families to identify informative single-nucleotide polymorphisms associated with mutant alleles. We performed five cycles of in vitro fertilization PGT-M in four couples with CMT1A, CMT2A, and CMT2S in CHA Fertility Center, Seoul Station. Results: From July 2020 through August 2021, five cycles of PGT-M with karyomapping in four cases with CMT1 and CMT2 were analyzed retrospectively. A total of 17 blastocysts were biopsied and 15 embryos were successfully diagnosed (88.2%). Ten out of 15 embryos were diagnosed as unaffected (66.7%). Five cycles of PGT-M resulted in four transfer cycles, in which four embryos were transferred. Three clinical pregnancies were achieved (75%) and the prenatal diagnosis by amniocentesis for all three women confirmed PGT-M of karyomapping. One woman delivered a healthy baby uneventfully and two pregnancies are currently ongoing. Conclusion: This is the first report in Korea on the application of karyomapping in PGT-M for CMT patients. This study shows that karyomapping is an efficient, reliable and accurate diagnostic method for PGT-M in various types of CMT diseases.