• Clinical and Experimental Reproductive Medicine
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• v.37 no.2
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• pp.99-113
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• 2010

Objective: It has been recently reported that very small stem cells with pluripotency are detected in murine and human. The purposes of this study are to confirm whether very small putative stem cells (VSPSCs), which have the proper characteristics of stem cells as well as the expression of stem cell markers, are detected in human endometrium. Methods: The endometrial cells of 5 women, which were obtained by endometrial biopsy, were cultured for 2 weeks and were confirmed for the expressions of alkaline phosphatase, OCT-4 and CXCR4 by immunochemistry. Subsequently VSPSCs were separated by percoll density gradient method and were cultured. Also VSPSCs and their derived cells were confirmed for the expressions of OCT-4 and CXCR4. Results: The colonies, which is composed with VSPSCs less than 3 ${\mu}m$ and the 5~15 ${\mu}m$ sized hyperchromatic round cells, were detected in the endometrium of all of women and showed the strong expressions of alkaline phosphatase, OCT-4 and CXCR4. In culture after the separation of VSPSCs by percoll, these cells showed the morphological and functional characteristics of stem cells; self-renewal, colony formation, embryoid body-like formation and differential plasticity. VSPSCs formed gradually the 5~15 ${\mu}m$ sized hyperchromatic round cells and the 10~20 ${\mu}m$ sized sphere-shaped cells by cell-to-cell aggregation or cell fusion. Then these cells differentiated the various cells including fibroblast-like cells, nerve-like cells and endothelium-like cells. VSPSCs and their derived cells often showed the strong expressions of OCT-4 and CXCR4. Conclusion: VSPSCs less than 3 ${\mu}m$ and their derived cells are detected in human endometrium and these cells have the proper characteristics of stem cells and the expressions of stem cell markers as alkaline phosphatase, OCT-4 and CXCR4.

• Journal of Science and Technology Studies
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• v.5 no.1 s.9
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• pp.93-123
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• 2005

This work has following two research goals. First, IVF treatments that have been recently going on in Korea are reexamined from the perspective of women's reproductive rights. Second, the intimate connection between IVF and therapeutic cloning research, in that remnant embryos and eggs that have been secured through IVF treatments have served as a main source of supply for therapeutic cloning research, has been emphasized. The fact that the influencing power of tradition on Korean families and women and IVF techniques eventually joined their hands in support of therapeutic cloning research is noted. Analysis of experiences of infertility by women in the realms of family, medical care during IVF treatment, and therapeutic cloning research that requires continuous supply of eggs leads to following conclusions. First, in the realm of family, infertile women were not only relegated to the status of abnormality but pressured to question their own womanhood. Under this circumstance, IVF treatment helped to reinforce the traditional concept of biological motherhood, thus categorizing married women giving birth to babies and married women who can't or refuses to do so to 'normal ones' and 'abnormal ones' respectively. Second, in the realm of medical care an infertile woman could rediscover her own body during the process of IVF treatment. By going through the processes of hormone treatment, implantation, conception, miscarriage, and so on, she could realize that her own body is understood in diverse ways to her, her family, and the medical profession. Third, in the realm of the state, IVF treatment that was serving as the main supplier of research materials for therapeutic cloning research has been able to avoid controversy in public discourses since the latter has emerged as a signifier of new national economic workhorse for the 21st century. As therapeutic cloning research went into high gear, the status of women as egg providers began to assume a political dimension. Women as egg providers are called upon to take on a paradoxical role as patriotic contributors to national economy on the one hand and as guardians of sacred 'life' on the other hand. The direction and progress of the research will depend on the ways that women comply, compromise, and/or resist the contradiction brought about by being assigned to assume these two identities: the one as a member of the nation requested to serve as a part of national economic development project, even though considered ineligible for financial recompense, and the other one as a guardian of sacred 'life,' even though she have to serve the research that is allowed to create a 'life' to destroy a 'life.'

• Journal of Life Science
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• v.30 no.8
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• pp.701-707
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• 2020

Gene editing technology using the clustered regularly interspaced short palindromic repeat (CRISPR) and the CRISPR associated protein (Cas)9 has been highly anticipated in developing breeding techniques. In this study, we discuss gene scissors as a tool for silkworm molecular breeding through analysis of Bombyx mori Kynurenine 3-Monooxygenase (BmKMO) gene editing using the CRISPR/Cas9 system and analysis of generational transmission through mutagenesis and selective crossing. The nucleotide sequence of the BmKMO gene was analyzed, and three guide RNAs (gRNAs) were prepared. Each synthesized gRNA was combined with Cas9 protein and then analyzed by T7 endonuclease I after introduction into the BM-N silkworm cell line. To edit the silkworm gene, K1P gRNA and Cas9 complexes were subsequently microinjected into the silkworm embryos; the hatching rate was 18% and the incidence of mutation was 60%. The gene mutation was verified in the heterozygous G0 generation, but no phenotypic change was observed. In homozygotes generated by self-crossing, a mutant phenotype was observed. These results suggest that silkworm molecular breeding using the CRISPR/Cas9 system is possible and could be an effective way of shortening the time required.

• Clinical and Experimental Reproductive Medicine
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• v.24 no.1
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• pp.119-133
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• 1997

In spite of much progress in vitro fertilization and embryo transfer (IVF-ET) program, the pregnancy rate remains at 20-30%, and the endometrial implantation rate per embryo transferred at 10-15%. As a result, about 90% of embryos may fail to implant to the endometrium, and many attempts such as optimization of follicular development, improvement of in vitro culture system including coculture, and micromanipulation of zona pellucida have been made to improve embryonic implantation after IVF-ET. Recently, several procedures of assisted hatching (AH) using micromanipulation have been introduced, and pregnancies and births have been obtained after AH. To develop and establish AH as an effective procedure to improve embryonic implantation, AH with partial zona dissection (PZD) was performed in 116 cycles of 89 infertile couples who had previous repeated failures of standard IVF-ET more than two times (Group I: 71 cycles in 54 patients), or who had implantation failure of embryos with good quality (Group II: 15 cycles in 13), or who had undergone AH without specific indication (Group III: 30 cycles in 22) from January, 1995 to Februry, 1996, and the outcomes of AH were analyzed according to pregnancy rate. The number of oocytes retrieved after controlled ovarian hyperstimulation (COH) was $9.9{\pm}7.1$ in Group I, $11.5{\pm}4.5$ in Group II, and $7.9{\pm}6.4$ in Group III. The number of embryos transferred after AH was $4.7{\pm}1.8$ in Group I, $5.3{\pm}1.3$ in Group II, and $3.5{\pm}2.4$ in Group III. The mean cumulative embryo score (CES) was $56.8{\pm}30.0$ in Group I, $76.1{\pm}35.9$ in Group II, and $38.5{\pm}29.9$ in Group III. The overall clinical pregnancy rate per cycle and per patient was 12.7% (9/71) and 16.7% (9/54) in Group I, 33.3% (5/15) and 38.5% (5/13) in Group II, and 6.7% (2/30) and 9.1% (2/22) in Group III, respectively. There were significant differences in the numbers of oocytes retrieved and embryos transferred, CES, and the clinical pregnancy rate per cycle among three groups. There was a significant inverse correlation between basal serum FSH level and CES, and no pregnancy occurred in patients with CES less than 20. In conclusion, AH of human embryos with PZD prior to ET has improved the implantation and pregnancy rates in IVF-ET patients with the past history of repeated failures, especially in spite of transfer of embryos with good quality, and AH will provide a range of novel techniques which may contribute much to effective management of infertile couples.

• Clinical and Experimental Reproductive Medicine
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• v.26 no.2
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• pp.137-148
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• 1999

Purpose of the present study was to find the optimal ovulation induction medicine for the maturation and development of immature oocytes and culture media for 2-cell embryos in the mouse model. ICR female mouse aged 6 to 8 weeks, were stimulated with 5 IU PMSG injection. At 47 to 50 hour post-PMSG injection, ovaries were dissected out and oocytes-cumulus complexes were punctured. The oocyte-cumulus complexes were cultured in media containing various ovulation induction medicine, CC, HMG and Metrodin for 18 hours. Female ICR mice were stimulated with 5 IU PMSG and 48 hours later were injected 5 IU of hCG, then female and male mice were mated. At 48 hour post-hCG injection, oviducts were dissected out and 2-cell embryos were flushed. The 2-cell embryos were cultured in various media, Ham's F-10 media of milli-Q water $(3^{\circ})$, Ham's F-10 media of HPLC (high performance liquid chromatography, Baxter) water, Medicult media, HTF (human tubal fluid) media for 96 hours. The results were as follows. 1. When the oocytes-cumulus complexes were cultured in $10^{-9}{\mu}g/ml{\sim}10^{-8}{\mu}g/ml$ of CC, those were suppressed in meiotic maturation $(28.2{\sim}33.7%)$. Whereas the oocytes-cumulus complexes were cultured in $10^{-7}{\mu}g/ml{\sim}10^{-4}{\mu}g/ml$, these were not effected in meiotic maturation $(54.5{\sim}72.7%)$. 2. When the oocytes-cumulus complexes were cultured in $10^{-4}{\mu}g/ml{\sim}10^{-1}{\mu}g/ml$ of Metrodin, those were suppressed in meiotic maturation $(35.7{\sim}41.5%)$. Meanwhile the oocytes-cumulus complexes were cultured in $10^{-7}{\mu}g/ml{\sim}10^{-5}{\mu}g/ml$, those were not effected in meiotic maturation $(54.2{\sim}70.3%)$. 3. When the oocytes-cumulus complexes were cultured in $10^{-5}{\mu}g/ml{\sim}10^{-4}{\mu}g/ml$ of HMG, those were suppressed in meiotic maturation $(48.2{\sim}50.4%)$. As being cultured in $10^{-7}{\mu}g/ml{\sim}10^{-6}{\mu}g/ml$, increased in meiotic maturation $(75.8{\sim}80.7%)$. 4. When the 2-cell embryos were cultured in Ham's F-10 media of milli-Q water $(3^{\circ})$, Ham's F-10 media of HPLC (high performance liquid chromatograpy, Baxter) water, Medicult media, HTF (human tubal fluid) media, developmental rates to blastocyst and hatching for 96 hour were 50.0%, 45.2%, 71.5% and 95.6%, respectively.

• Korean Journal of Environmental Biology
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• v.38 no.2
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• pp.271-277
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• 2020

Toxicities to many organs caused by humidifier disinfectants have been reported. Recently, humidifier disinfectants have been reported to cause cardiovascular, embryonic, and hepatic toxicities. This study was designed to investigate the toxic mechanism of humidifier disinfectants and compare toxicity in a cellular model and a zebrafish animal model. Because brain toxicity and skin toxicity have been less studied than other organs, we evaluated toxicity in a human dermal cell line and zebrafish under various concentrations of humidifier disinfectants that included polyhexamethyleneguanidine phosphate (PHMG), oligo-[2-(2-ethoxy)-ethoxyethyl-guanidinium-chloride] (PGH) and methylchloroisothiazolinone/methylisothiazolinone (CMIT/MIT). A human dermal fibroblast cell line was treated with disinfectants (0, 2, 4, 6, 8, and 16 mg L^-1) to compare their cytotoxicity. The fewest PHMG-treated cells survived (up to 33%), while 49% and 40% of the PGH- and CMIT/MIT-treated cells, respectively, survived. The quantification of oxidized species in the media revealed that the PHMG-treated cells had the highest MDA content of around 28 nM, while the PGH- and CMIT/MIT-treated cells had 13 and 21 nM MDA, respectively. As for brain toxicity, treatment of the zebrafish tank water with CMIT/MIT (final 40 mg L^-1) for 30 min resulted in a 17-fold higher production of reactive oxygen species (ROS) than in the control. Treatment with PGH or PHMG (final 40 mg L^-1) resulted in 15- and 11-fold higher production, respectively. The humidifier disinfectants (PHMG, PGH, and CMIT/MIT) showed severe dermal cell toxicity and brain toxicity. These toxicities may be relevant factors in understanding why some children have language disorders, motor delays, and developmental delays from exposure to humidifier disinfectants.

• Korean Journal of Veterinary Research
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• v.42 no.2
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• pp.137-146
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• 2002

The immunohistochemical localization of the nerve growth factor (NGF), glial fibrillary acidic protein (GFAP) and ciliary neurotrophic factor (CNIF) in the developing Mongolian gerbil forebrain was investigated by the immunohistochemical and electron microscopy methods. Generally, the NGF specifically recognizes the neurons, the GFAP does the glia, and the CNIF does the motor neurons. This study demonstrates the location of the NGF, GFAP and CNTF in the developing Mongolian gerbil from the embryonic days 17 (E17) to the postnatal weeks 3 (PNW 3). The NGF was localized at E19 in the olfactocy bulb and the cerebral cortex, expanded to the hippocampus, and the diagonal bond from the late prenatal period to PNW 3. GFAP was observed in the lateral ventricle and the third ventricle at E17, projected into the cerebral cortex at E19. The GFAP was observed to have the largest numbers in several parts of the forebrain at the postnatal days 2 (PND2), while the most numerous CNTF was observed at PNW 2. The CNTF-IR cells were observed only in the postnatal days and were found in the olfactory bulb, cerebral cortex both neuron and neuroglia at PND3. Electron microscopy showed that the NGF, GFAP and CNTF were not related to any connections with any particular subcellular structure. These results suggest that NGF, GFAP and CNTF be related to the neuron and neuroglia at the prenatal and postnatal stages in the developing Mongolian gerbil.

• Journal of the Korea Academia-Industrial cooperation Society
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• v.19 no.9
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• pp.116-123
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• 2018

The sperm quality is determined by the kinetic characteristics and acrosome integrity of the sperm. In the previous studies, analysis of semen quality had large errors because those experiments by using microscope had been conducted by people. In recent years, the molecular biological methods have been newly developed to complement the previous techniques. The ZAR1 gene is known to be a gene that affects early embryonic development in vertebrates, but there is no study of the association with semen. In this study, we analyzed the association between the kinetic characteristics and ZAR1 single nucleotide polymorphism (SNP) genotype. To detect the SNPs, we performed sequencing using genomic DNA from the whole bloods of Duroc pigs. We identified an SNP in the ZAR1 gene g.2540T>C. ZAR1 SNP genotypeing in 105 pigs revealed that the major and minor alleles were T and C, respectively. After we analyzed the association between the kinetic characteristics of sperm and the ZAR1 SNP genotype, we found a significant association in MOT (p<0.01), VSL (p<0.05) of the kinetic characteristics in the Duroc boars. It was confirmed that the boars with T allele were lower in MOT and VSL than C allele. Therefore, pigs with C allele are judged to be better at the MOT and VSL of semen. Based on these results, ZAR1 SNP genotyping may be a useful molecular biomarker to improve semen quality by applying molecular breeding technology.

• Clinical and Experimental Pediatrics
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• v.48 no.2
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• pp.186-190
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• 2005

Purpose : The purpose of this study was to evaluate the responses and toxicities of risk-adapted chemotherapy in pediatric intracranial germ cell tumors(IC-GCT). Methods : Fourteen patients who were diagnosed as IC-GCT from October 2002 to December 2003 received chemotherapy as an initial treatment modality. The low risk(LR) group was defined as follows : Pure germinoma and normal AFP level. Beta-hCG level 50 mIU/mL or less. The others belonged to the high risk(HR) group. Chemotherapy was composed of cisplatin, cyclophosphamide, etoposide and vincristine. Double doses of cisplatin and cyclophosphamide was used in HR patients. Results : Pathologic confirmation was done in all but one. Median age at diagnosis was 11.6 yr (1.2-18.7 yr), and nine patients belonged to the HR group. Tumor markers were normalized after chemotherapy in all patients whose tumor markers had been elevated. Four LR patients(80 percent) and seven HR patients(77.8 percent) showed complete response(CR) at the end of chemotherapy. An additional two of the three patients with partial response(PR) achieved CR after radiation therapy (RT), and the remaining one relapsed before RT. Four LR and all HR patients experienced infectious episodes that required hospitalization. Four of the nine HR patients(44.4 percent) suffered from tinnitus, three of whom developed sensorineural hearing loss. All but one are surviving, event-free, with a median follow-up of 13.9 mo(8.1-22.3 mo). Conclusion : Risk-adapted cisplatin-based chemotherapy was effective even in HR patients, but regimen modification seems to be necessary to avoid an unacceptably high toxicity rate.

• Development and Reproduction
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• v.13 no.4
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• pp.353-362
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• 2009

GnRH and its receptor are known to express locally in the ovary and to regulate the ovarian function by affecting on granulosa and lutein cells. It has been reported that GnRH directly causes apoptosis in the granulosa and lutein cells of the ovary. However, whether the apoptosis of the cells by GnRH is recovered by FSH as an anti-apoptotic factor is not yet known. In this study, we evaluated the apoptosis and the production of progesterone $(P_4)$ and estradiol $(E_2)$ after treatment with 5, 50, and 100 ng/$m\ell$ GnRH and 1 IU/ml FSH in the granulosa-lutein cells that are obtained during oocyte-retrieval for IVF-ET. Results of DNA fragment analysis and TUNEL assay demonstrated that DNA fragmentation and the rate of apoptotic cells were increased in a dose-dependent manner showing a significant increase in the cells treated with 100 ng/$m\ell$ GnRH. In addition, we found that FSH suppresses the apoptosis of the cells induced by GnRH. In the results of chemiluminescence assay for $P_4$ and $E_2$, $P_4$ production was decreased by GnRH treatment, whereas $E_2$ production was not changed. We also demonstrated that FSH inhibits the suppressive effect of GnRH on $P_4$ production as the result of apoptosis. The present results suggest that GnRH agonist using in ovarian hyperstimulation protocol might induce the dysfunction of the ovary, but its function could be recovered by FSH. These results also will be expected to use as the basic data to elucidate the physiological role of GnRH and to develop new ovarian hyperstimulation protocols for IVF-ET.