• Animal Bioscience
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• v.36 no.12
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• pp.1905-1917
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• 2023

Objective: Nanog homeobox (NANOG) is a core transcription factor that contributes to pluripotency along with octamer binding transcription factor-4 (OCT4) and sex determining region-Y box-2 (SOX2). It is an epiblast lineage marker in mammalian pre-implantation embryos and exhibits a species-specific expression pattern. Therefore, it is important to understand the lineage of NANOG, the trophectoderm, and the primitive endoderm in the pig embryo. Methods: A loss- and gain-of-function analysis was done to determine the role of NANOG in lineage specification in parthenogenetic porcine blastocysts. We analyzed the relationship between NANOG and pluripotent core transcription factors and other lineage makers. Results: In NANOG-null late blastocysts, OCT4-, SOX2-, and SOX17-positive cells were decreased, whereas GATA binding protein 6 (GATA6)-positive cells were increased. Quantitative real-time polymerase chain reaction revealed that the expression of SOX2 was decreased in NANOG-null blastocysts, whereas that of primitive endoderm makers, except SOX17, was increased. In NANOG-overexpressing blastocysts, caudal type homeobox 2 (CDX2-), SOX17-, and GATA6-positive cells were decreased. The results indicated that the expression of primitive endoderm markers and trophectoderm-related genes was decreased. Conclusion: Taken together, the results demonstrate that NANOG is involved in the epiblast and primitive endoderm differentiation and is essential for maintaining pluripotency within the epiblast.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.23
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• pp.10255-10262
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• 2015

High risk human papillomavirus (HR-HPV) E2 proteins play roles in transcriptional regulation and are commonly functionally disrupted when the HPV genome integrates into host chromosomes. Some 15-40% of cancer cases, however, contain an intact E2 gene or episomal HPV. In these cases, polymorphism of the E2 gene might be involved. This study aimed to determine polymorphisms of the E2 gene in episomal HPV16 detected in high grade squamous intraepithelial lesions and squamous cell carcinomas and altered functions compared to the E2 prototype. The E2 gene was amplified and sequenced. Two expression vectors containing E2 gene polymorphisms were constructed and transfected in SiHa and C33A cells, then E6 gene as well as Il-10 and TNF-${\alpha}$ expression was determined by quantitative RT-PCR. Expression vectors and reporter vectors containing the HPV16 long control region (LCR) were co-transfected and transcriptional activity was determined. The results showed that a total of 32 nucleotides and 23 amino acids were changed in all 20 cases of study, found in the transactivation (TA) domain, hinge (H) region and DNA binding (DB) domain with 14, 5 and 13 nucleotide positions. They mostly caused amino acid change. The expressing vectors containing different E2 gene polymorphisms showed E6 mRNA suppression, TNF-${\alpha}$ mRNA suppression and IL-10 induction but no statistically significant differences when compared to the E2 prototype. Moreover, promoter activity in HPV16 LCR was not affected by E2 protein with different gene polymorphisms, in contrast to nucleotide variations in LCR that showed an effect on transcription activity. These results demonstrated that E2 gene polymorphisms of episomal HPV16 did not affect transcriptional regulation and suggested that nucleotide variation as well as epigenetic modification of the LCR might play a role in inducing malignant transformation of cells containing episomal HPV16.

• Journal of Korean Ophthalmic Optics Society
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• v.12 no.2
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• pp.67-78
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• 2007

The eye irritant test of lens washing agent, CLIWELL$^{TM}$ was conducted using Draize methods according to KFDA Guidelines. In addition, to test the potential toxicity of test articles, the ratio of inflammatory cells and non-inflammatory epitheloid cells was also observed using smear cytology methods against ocular discharge. At sacrifice, the histopathological changes on Cornea, Iris, Retina and Sclera were also observed in all animals. Slight irritancy of cornea and conjunctiva was observed at 1, 2 and 3 days after dropping in non-washing group. The MIOI of these points are detected as 4.50, 1.67 and 0.67, respectively. In washing group, slight irritancy of cornea and conjunctiva were observed at 1 and 2 days after dropping with MIOI as 0.67, respectively. Therefore, CLIWELL$^{TM}$ was considered as non-irritating materials because the MIOI is detected below 5.00 throughout the whole experimental periods in both washing and non-washing groups and the IAOI was also detected as 4.50. Except for significant increase of the inflammatory cell ratios in ocular discharge at 1 day after dropping of non-washing group compared to that non-treated intact eyes, no meaningful changes on smear cytology of ocular discharges are observed in this study. In addition, no abnormal histopathological changes on the cornea, iris, retina and sclera were also detected in CLIWELL$^{TM}$ dropping group compared to that of non-treated intact eyes.

• Tuberculosis and Respiratory Diseases
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• v.46 no.2
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• pp.273-280
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• 1999

This is a report of gastric metastases secondary from a primary small cell carcinoma of the lung in two men. Blood-borne metastatic involvement of the stomach by cancer is a rare entity. According to the reports in the literature the prevalence of metastasis to the stomach occurs in 0.4% and the most common cell type of the primary lung carcinoma is large cell type(3.7%) followed by adenocarcinoma(2.4%), small cell carcinoma(1.7%) and squamous cell carcinoma(0.7%). The most common tumors that spread to the stomach through the blood stream are malignant melanoma, breast carcinoma and lung carcinoma. Most of the gastrointestinal tract metastases had no specific symptoms because of its submucosal involvement. The prognosis was poor and the mean survival period from the onset of symptoms was 49 days. The first patient was a 56-year-old man who had primary lung carcinoma with brain metastasis. Gastroscopic findings showed two elevated mass lesions in the anterior wall of the mid body with central ulcer and the posterior wall of the fundus with intact surface mucosa. Pathologic examination of stomach tissue revealed small cell type tumor cells infiltrate in the stomach wall segmentally without destruction of the glands. The second patient was a 67-year-old man who had no other evidence of the distant metastasis. Gastroscopic findings showed a huge, oval shaped, ulcerofungating mass with deep penetrating central ulcer coated with dirty exudate in the anterior wall from mid to upper body of the stomach, and thickened elevated rugal folds in the posterior wall of the fundus. Pathologic examination of stomach tissues revealed the small cell type tumor cells showing small smudged nucleus infiltrate into the mucosa of the stomach and the architecture of mucosa intact. We report the two cases of metastatic gastric cancer from the primary small cell lung carcinoma with the literature review.

• Journal of Korean Ophthalmic Optics Society
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• v.11 no.4
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• pp.299-310
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• 2006

The eye irritant test of lens washing agent, $SOLOCARE^{TM}$ was conducted using Draize methods according to KFDA Guidelines 1999-61. In addition, to test the potential toxicity of test articles the ratio of inflammatory cells and non-inflammatory epitheloid cells were also observed using smear cytology methods against ocular discharge. At sacrifice, the histopathological changes on Cornea, Iris, Retina and Sclera were also observed in all animals. Slight (1~2 degrees) irritancy of cornea and conjunctiva were observed at 1, 2 and 3 days after dropping in non-washing group. The MIOI of these points are detected as 4.33, 3.33 and 2.00, respectively. In washing group, slight irritancy of cornea and conjunctiva were restricted to 1 and 2 days after dropping with MIOI as 3.00 and 1.33, respectively. Therefore, $SOLOCARE^{TM}$ was also considered as non-irritating materials because the MIOI is detected below 5.00 throughout the whole experimental periods in both washing and non-washing groups and the IAOI is also detected as 4.33 (1 day after dropping). No meaningful changes on smear cytology of ocular discharges are observed in this study compared to that of non-treated intact eyes. In addition, no abnormal histopathological changes on the cornea, iris, retina and sclera were also detected in $SOLOCARE^{TM}$ dropping group compared to that of non-treated intact eyes.

• The Korean Journal of Physiology and Pharmacology
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• v.21 no.6
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• pp.591-598
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• 2017

Propofol is known to cause vasorelaxation of several systemic vascular beds. However, its effect on the pulmonary vasculature remains controversial. In the present study, we investigated the effects of propofol on human pulmonary arteries obtained from patients who had undergone surgery. Arterial rings were mounted in a Multi-Myograph system for measurement of isometric forces. U46619 was used to induce sustained contraction of the intrapulmonary arteries, and propofol was then applied (in increments from $10-300{\mu}m$). Arteries denuded of endothelium, preincubated or not with indomethacin, were used to investigate the effects of propofol on isolated arteries. Propofol exhibited a bifunctional effect on isolated human pulmonary arteries contracted by U46619, evoking constriction at low concentrations ($10-100{\mu}m$) followed by secondary relaxation (at $100-300{\mu}m$). The extent of constriction induced by propofol was higher in an endothelium-denuded group than in an endothelium-intact group. Preincubation with indomethacin abolished constriction and potentiated relaxation. The maximal relaxation was greater in the endothelium-intact than the endothelium-denuded group. Propofol also suppressed $CaCl_2$-induced constriction in the 60 mM $K^+$-containing $Ca^{2+}$-free solution in a dose-dependent manner. Fluorescent imaging of $Ca^{2+}$ using fluo-4 showed that a 10 min incubation with propofol ($10-300{\mu}m$) inhibited the $Ca^{2+}$ influx into human pulmonary arterial smooth muscle cells induced by a 60 mM $K^+$-containing $Ca^{2+}$-free solution. In conclusion, propofol-induced arterial constriction appears to involve prostaglandin production by cyclooxygenase in pulmonary artery smooth muscle cells and the relaxation depends in part on endothelial function, principally on the inhibition of calcium influx through L-type voltage-operated calcium channels.

• The Korean Journal of Physiology and Pharmacology
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• v.22 no.2
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• pp.215-223
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• 2018

Intracellular $Ca^{2+}$ mobilization is closely linked with the initiation of salivary secretion in parotid acinar cells. Reactive oxygen species (ROS) are known to be related to a variety of oxidative stress-induced cellular disorders and believed to be involved in salivary impairments. In this study, we investigated the underlying mechanism of hydrogen peroxide ($H_2O_2$) on cytosolic $Ca^{2+}$ accumulation in mouse parotid acinar cells. Intracellular $Ca^{2+}$ levels were slowly elevated when $1mM\;H_2O_2$ was perfused in the presence of normal extracellular $Ca^{2+}$. In a $Ca^{2+}-free$ medium, $1mM\;H_2O_2$ still enhanced the intracellular $Ca^{2+}$ level. $Ca^{2+}$ entry tested using manganese quenching technique was not affected by perfusion of $1mM\;H_2O_2$. On the other hand, $10mM\;H_2O_2$ induced more rapid $Ca^{2+}$ accumulation and facilitated $Ca^{2+}$ entry from extracellular fluid. $Ca^{2+}$ refill into intracellular $Ca^{2+}$ store and inositol 1,4,5-trisphosphate ($1{\mu}M$)-induced $Ca^{2+}$ release from $Ca^{2+}$ store was not affected by $1mM\;H_2O_2$ in permeabilized cells. $Ca^{2+}$ efflux through plasma membrane $Ca^{2+}-ATPase$ (PMCA) was markedly blocked by $1mM\;H_2O_2$ in thapsigargin-treated intact acinar cells. Antioxidants, either catalase or dithiothreitol, completely protected $H_2O_2-induced$ $Ca^{2+}$ accumulation through PMCA inactivation. From the above results, we suggest that excessive production of $H_2O_2$ under pathological conditions may lead to cytosolic $Ca^{2+}$ accumulation and that the primary mechanism of $H_2O_2-induced$ $Ca^{2+}$ accumulation is likely to inhibit $Ca^{2+}$ efflux through PMCA rather than mobilize $Ca^{2+}$ ions from extracellular medium or intracellular stores in mouse parotid acinar cells.

• Applied Microscopy
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• v.38 no.3
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• pp.205-212
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• 2008

In this study morphological changes of the testes of golden hamsters treated with 6-aminonicotinamide (6-AN, 10 mg/kg body weight) in every two days were investigated using light and transmission electron microscopes. After the 7th injection of 6-AN, body weights of hamsters were significantly reduced, and the weight of testes were markedly reduced in the group of hamsters after 5th injections. Degeneration of seminiferous epithelium appeared first in the group receiving 5th injections, which were followed by severe degenerations after the 9th injection. In the degenerated seminiferous epithelium, deep vacuolization, and destruction of spermatogenic cells and Sertoli cells were also oberved. Multinuclear giant cells were also observed in the lumen of destructed seminiferous epithelium. But there were no edematous changes in the interstitial tissue, and the Leydig cells were found to be relatively intact. Therefore, these results show that 6-AN trigger severe morphological alteration of the spermatogenic cells and Sertoli cells, however Leydig cells are unaltered by 6-AN.

• Journal of Animal Science and Technology
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• v.46 no.4
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• pp.571-584
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• 2004

It is known widely that granulosa cell apoptosis leads follicular atresia and macrophages exert their effects directly and/or indirectly from the initiation to the completion of follicular atresia by phagocytosis of apoptotic bodies and secretion of various cytokines. However, the site of initiation, propagation routes and the elimination methods of apoptotic bodies, and the time and methods of penetration of macrophages into the follicles are not known completely. Using pig(Yorkshire-breed) ovary, immunohistochemical studies with TUNEL for apoptotic bodies and pig macrophage monoclonal antibody 4E9 for macrophages, and light and transmission electron microscopic observations were performed. In the pig, follicular atresia began with the granulosa cell apoptosis, and the apoptosis of theca intema cells occured at the same time. The apoptosis of granulosa cells initiated randomly within the granulosa cell layer and propagated rapidly into the whole layer. Ultrastructura1ly, apoptotic granulosa cells showed characteristic pyknotic and deformed nucleus and intracytoplasmic vesicles. Apoptotic bodies were eliminated by intact granulosa cells and macrophages. Intact granulosa cells ingested apoptotic bodies transiently, soon after they fell into the apoptosis. Finally, apoptotic bodies and degenerated oocyte were phagocytosed by macrophages. Macrophages entered the ovarian follicle at the time of initiation of granulosa cell apoptosis, and migrated with the progression of apoptosis. By elimination of theca cells, macrophages contributed the completion of follicular atresia These results will provide valuable informations on the study of the interrelation between macrophage and ovarian follicular atresia.

• Korean Journal of Poultry Science
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• v.28 no.2
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• pp.135-142
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• 2001

A novel sterategy has been established to determine the origin of the Primordial Germ Cells (PGCs) in avian embryos directly and the developmental fate of the PGCs for the application to Poultry biotechnology. Cells were removed from 1) the centre of area pellucida, 2) the outer of area pellucida and 3) the area opaca of the stage X blastoderm (Eyal-Giladi & Kochav, 1976). When the cells were removed from the centre of area pellucida, the mean number of circulating PGCs in blood was significantly decreased in the embryo at stage 15 (Hamburger & Hamilton, 1951) as compared to intact embryos. When the cells were replenished with donor cells, no reduction in the PGCs number was observed. The removal of cells at the outer of area pellucida or at the area opaca had no effect on the number of PGCs. In case, another set of the manipulated embryos were cultured ex vivo to the hatching and reared to the sexual maturity, the absence of germ cells and degeneration of seminiferous tubules was observed in resulting chickens derived from the blastoderm in which the cells were removed from the centre of the area pellucida. It was concluded that the avian Primordial Germ cells are originated at the center of area pellucida. Developmental ability of the cells to differentiate into somatic cells and germ cells in chimeras were analyzed. Somatic chimerism was detected as black feather attributed from donor cells. Molecular identification by use of female - specific DNA was performed. It was confirmed that the donor cells could be differentiated into chimeric body and erythrocytes. Donor cells retained the ability to differentiate into germline in chimeric gonads. More than 70% of the generated chimeras transmitted donor derived gametes to their offspring indicating that the cells at the center of area pellucida had the high ability to differentiate into germ cells. A molecular technique to identify germline chimerism has been developed by use of gene scan analysis. Strain specific DNA fragments were amplified by the method. It would be greatly contributed for the detection of germline chimerism. Mixed- sex chimeras which contained both male and female cells were produced to investigate the developmental fate of male and female cells in ovary and testes. The sex combinations of donor and recipient of the resulting chimeras were following 4 pairs; (1) chimeras (ZZ/ZZ) produced by a male donor (ZZ) and a male recipient (ZZ), (2) chimeras (ZW/ZW) produced by a female donor (ZW) and a female recipient (ZW), (3) chimeras (ZZ/ZW) Produce by a male donor (ZZ) and a female recipient (ZW), (4) chimeras (ZW/ZZ) produced by a female donor (ZW) and a male recipient (ZZ). It was found that genetically male avian germ cells could differentiate into functional ova and that genetically female germ cells can differentiate into functional spermatozoa in the gonad of the mixed- sex chimeras. An ability for introduction of exogenous DNA into the PGCs from stage X blastoderms were analyzed. Two reporter genes, SV-$\beta$gal and RSV-GFP, were introduced into the PGCs. Expression of bacterial/gal was improved by complexing DNA with liposome detectedcc in 75% of embryos at 3 days embryos. At the embryos incubated for 1 day, expression of the GFP was observed all the embryos. At day 3 of incubation, GFP was detected in about 70% of the manipulated embryos. In case of GFP, expression of the transgene was detected in 30 %e of the manipulated embryos. These results suggested that the cells is one of the most promising vectors for transgenesis. The established strategy should be very powerfull for application to poultry biotechnology.