• Journal of Life Science
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• v.29 no.10
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• pp.1159-1163
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• 2019

Although there is a growing interest in male menopause, a phenomenon associated with male hormone depletion, current kits using antibodies to quantify male hormones are expensive. In this study, we constructed an in vitro system for verifying the activity or concentration of male steroid hormones using a transcriptional activity test. A reporter plasmid, pGL2-Neo-ARE-AdE1BTATA, which reacts to testosterone, was constructed. In this plasmid, the ARE-AdE1bTATA sequences can be bounded by the testosterone - androgen receptor complex to express luciferase as a reporter. Then, a stable transfection was performed on the human prostate cancer cell line, LNcap-LN3. The constructed LNcap-LN3/pGL2-Neo-ARE-AdE1BTATA testosterone compound detection (TCD) system showed quantitatively proportional luciferase activities to concentrations of $10^{-13}$ to $10^{-8}M$ of standard testosterone. The established in vitro TCD system will contribute to the development of materials for health/functional foods and drugs as it will be possible to search en masse for testosterone-like or testosterone-inhibiting substances derived from natural materials.

• Journal of Physiology & Pathology in Korean Medicine
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• v.18 no.3
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• pp.740-746
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• 2004

In order to identify the anti-inflammatory and analgesic properties of natural herbal extracts, widely used in the Korean traditional medicine, an in vitro screening system was designed using pGL3, a luciferase reporter vector, and the tumor necrosis factor (TNF)-α and cyclooxygenase (COX)-II as target genes. The promoter regions of each gene was generated by PCR using the human chromosome as template DNA, and inserted into pGL3 vector with Kpnl and Hindlll. The final construct was transfected into human myleomonocytic leukemia cells (U937) that could be differentiated and activated by phorbol 12-myristate 13-acetate (PMA) or lipopolysaccharide (LPS). Using this system, we tested the anti-inflammatory and analgesic effects of several herbal extracts being regarded to have the medicinal effects of diminishing the body heat and complementing Qi. The well-known chemicals of PD98059 and berberine chloride were used as controls of the transcriptional inhibitors of TNF-α and COX-II, respectively. Among them, Salvia miltiorrhiza (Dan-Sam) was found to exhibit the significant medicinal properties of anti-inflammatory and analgesic effects.

• Development and Reproduction
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• v.14 no.2
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• pp.123-129
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• 2010

Embryonic stem (ES) cells can self-renew maintaining the undifferentiated state. Self-renewal requires many factors such as OCT4, SOX2, and NANOG. It is previously known that OCT4 and SOX2 can bind to NANOG promoter and support Nanog gene expression in mouse ES cells by the detailed studies using the mouse Nanog promoter. Here, we constructed serial deletion mutant promoter-reporter constructs to investigate the human Nanog gene promoter in detail. The highest promoter activity was obtained in the 0.6 kb (-253/+365) promoter-reporter construct which includes the binding sites of OCT4 and SOX2. To further confirm contribution of OCT4 and SOX2 in Nanog gene expression, we introduced site- directed mutation(s) in the OCT4 and/or SOX2 binding sites of the human Nanog promoter 0.6 kb (-253/+365) and checked the influence of the mutation on the promoter activity using human EC cell line NCCIT. Mutation either in OCT4 binding site or SOX2 binding site significantly reduced the activity of Nanog promoter which directly confirmed that OCT4 and SOX2 binding is essential in human Nanog gene expression.

• Reproductive and Developmental Biology
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• v.30 no.2
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• pp.81-85
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• 2006

This study was conducted to examine the effect of IRES controlled reporter gene on screening and production of recombinant human erythropoietin (EPO) proteins from cultured CHO cells. The cDNA was cloned for EPO from human liver cDNA Using site-directed mutagenesis, we generated recombinant human EPO (rhEPO) with two additional N-glycosylations (Novel erythropoiesis-stimulating protein: NESP). Wild type hEPO and NESP were cloned into expression vectors with GFP reporter gene under regulatory control of CMV promoter and IRES so that the vectors could express both rhEPO and GFP. The expression vectors were transfected to cultured CHO-K1 cells. Under microscopy, expression of GFP was visible. Using supernatant of the culture, ELISA assay, immunocytochemistry and in vitro assay using EPO dependant cell line were performed to estimate biological activity to compare the production characteristics (secretion levels, etc.) between rhEPO and NESP. The activity of NESP protein, obtained by mutagenesis, was described and compared with its rhEPO counterpart produced under same conditions. Although NESP had less secretion level in CHO cell line, the biological activity of NESP was greater than that of rhEPO. These results are consistent with previous researches. We also demonstrated that rhEPO and GFP proteins expressed simultaneously from transfected CHO cell line. Therefore we conclude that use of GFP reporter gene under IRES control could be used to screen and produce rhEPO in cultured CHO cells.

• Korean journal of communication and information
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• v.26
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• pp.45-82
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• 2004

The purpose of this study is to examine the concept of civic journalism and the contents of Korean Internet newspapers that might reflect the possibilities of this new medium for civic journalism practices. This study examined how far and deep civic journalism practices have extended into Korean Internet newspapers as journalism's new tradition. More specifically, this study analyzed news articles of Korean Internet newspapers to uncover any differences among civic journalism Internet newspapers with different citizen reporter systems. The composite measure based upon ten elements of civic journalism practices was used as indicator of civic journalism practices. To obtain systematic data on news offered by Korean Internet newspapers on the World Wide Web, four major Internet newspapers, including Ohmynews, Ngotimes, Netpinion and Pressian were examined by a content analysis in April and May of 2003. Findings of this study reveal that many Korean newspapers do not fully exploit the opportunities and advantages offered by the new medium for civic journalism practices in online environments. Both aggregate and individual level of analysis for the civic journalism index reveal that there are some differences between non-civic journalism and civic journalism Internet newspapers using citizen reporter systems. However, overall performances of civic journalism Internet newspapers are not good enough to support the argument that civic journalism is well practiced in Korean Internet newspapers. Nonetheless, it would not be fair to conclude that Korean Internet newspapers have totally ignored the Internet's potential to increase the civic journalism performance in online environments.

• The Korean Journal of Emergency Medical Services
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• v.11 no.2
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• pp.51-66
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• 2007

Purpose: With a view to providing basic data to develop cardiopulmonary resuscitation education suitable for elementary students, the cardiopulmonary resuscitation education was conducted to grasp students' knowledge, skills accuracy and the attitude change before and after the education. Methods: Convenience sampling was made on fourth and fifth graders(total-35 students) of S elementary school located in K city, Chungcheongnam-do, and this was a pre-experiment research designed before and after choosing a single group. In terms of methods, specifically we, researchers ; 1) Handed out questionnaires to students directly to make them fill in firsthand and collected the questionnaires. 2) Utilized PPT materials based on 2005 AHA guideline and DVD materials of AHA, to give students theoretical education of cardiopulmonary resuscitation. We used Anne/SkillReporter$^{(R)}$ torso produced by Leardal Inc, and Little Anne to conduct practical education individually. 3) Asked students to give Anne/SkillReporter$^{(R)}$ torso cardiopulmonary resuscitation five times with the ratio of 30 : 2, and then one of researchers filled in the evaluation sheet individually. 4) Evaluated the accuracy of students' ability to perform the resuscitation based on the record of Anne/SkillReporter$^{(R)}$ integrated printer(which was the objective tool to grasp students' skills accuracy). 5) Gave out questionnaires to make students fill them in and then collected them. after completing the practical evaluation. Results: 1) In case of the attitude about cardiopulmonary resuscitation, Students' confidency rose from 19.28%(before the education) to 93.57(after the education)- which is a positive change. 2) As the result of the education, some elementary students scored 11 points (full score-16 points), up from 5 points before the education, in terms of the knowledge about cardiopulmonary resuscitation. The average point also reached 13.14 points(after the education), jump from 8.37(before the education), which was the rise of 29.8%. 3) When it comes to the practical performance, the skills accuracy was 80.93% on average, and the calculation method was as follows: total items were 16, and each item was marked form 0 to 2 points, meaning the full score was 32 points. The minimum score was 19 points and the maximum was 32($M{\pm}SD=25.90{\pm}2.88$), which was calculated based on percentage. 4) Regarding skills accuracy, respiration accuracy(%)($M{\pm}SD=30.20{\pm}27.16$) was higher than pressure accuracy(%) ($M{\pm}SD=15.34{\pm}25.27$). Conclusion: The result showed that students' attitude on cardiopulmonary resuscitation changed positively. and meaningful difference(p = .00) existed in the change of students' knowledge. In terms of skills accuracy. chest compression and airway control showed high accuracy, but the result of Anne/SkillReporter$^{(R)}$ performance showed that the accuracy of chest compression was lower than that of mouth-to-mouth resuscitation.

• Journal of Periodontal and Implant Science
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• v.36 no.4
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• pp.839-847
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• 2006

The aim of this study is to monitor reporter gene expression under osteocalcin gene promoter, using a real-time molecular imaging system, as tool to investigate osteoblast differentiation. The promoter region of mouse osteocalcin gene 2 (mOG2), the best-characterized osteoblast-specific gene, was inserted in promoterless luciferase reporter vector. Expression of reporter gene was confirmed and relationship between the reporter gene expression and osteoblastic differentiation was evaluated. Gene expression according to osteoblstic differentiation on biomaterials, utilizing a real-time molecular imaging system, was monitored. Luciferase was expressed at the only cells transduced with pGL4/mOGP and the level of expression was statistically higher at cells cultured in mineralization medium than cells in growth medium. CCCD camera detected the luciferase expression and was visible differentiation-dependent intensity of luminescence. The cells produced osteocalcin with time-dependent increment in BMP-2 treated cells and there was difference between BMP-2 treated cells and untreated cells at 14days. There was difference at the level of luciferase expression under pGL4/mOGP between BMP-2 treated cells and untreated cells at 3days. CCCD camera detected the luciferase expression at cells transduced with pGL4/mOGP on Ti disc and was visible differentiation-dependent intensity of luminescence This study shows that 1) expression of luciferase is regulated by the mouse OC promoter, 2) the CCCD detection system is a reliable quantitative gene detection tool for the osteoblast differentiation, 3) the dynamics of mouse OC promoter regulation during osteoblast differentiation is achieved in real time and quantitatively on biomaterial. The present system is a very reliable system for monitoring of osteoblast differentiation in real time and may be used for monitoring the effects of growth factors, drug, cytokines and biomaterials on osteoblast differentiation in animal.

• Molecules and Cells
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• v.41 no.5
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• pp.423-435
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• 2018

This investigation was aimed at working out the combined role of lncRNA H19, miR-29b and Wnt signaling in the development of colorectal cancer (CRC). In the aggregate, 185 CRC tissues and corresponding para-carcinoma tissues were gathered. The human CRC cell lines (i.e. HT29, HCT116, SW480 and SW620) and normal colorectal mucosa cell line (NCM460) were also purchased. Si-H19, si-NC, miR-29b-3p mimics, miR-29b-3p inhibitor, si-PGRN and negative control (NC) were, respectively, transfected into the CRC cells. Luciferase reporter plasmids were prepared to evaluate the transduction activity of $Wnt/{\beta}-catenin$ signaling pathway, and dual-luciferase reporter gene assay was arranged to confirm the targeted relationship between H19 and miR-29b-3p, as well as between miR-29b-3p and PGRN. Finally, the proliferative and invasive capacities of CRC cells were appraised through transwell, MTT and scratch assays. As a result, overexpressed H19 and down-expressed miR-29b-3p displayed close associations with the CRC patients' poor prognosis (P < 0.05). Besides, transfection with si-H19, miR-29b-3p mimic or si-PGRN were correlated with elevated E-cadherin expression, decreased snail and vimentin expressions, as well as less-motivated cell proliferation and cell metastasis (P < 0.05). Moreover, H19 was verified to directly target miR-29b-3p based on the luciferase reporter gene assay (P < 0.05), and miR-29b-3p also bound to PGRN in a direct manner (P < 0.05). Finally, addition of LiCl ($Wnt/{\beta}-catenin$ pathway activator) or XAV93920 ($Wnt/{\beta}-catenin$ pathway inhibitor) would cause remarkably altered E-cadherin, c-Myc, vimentin and snail expressions, as well as significantly changed transcriptional activity of ${\beta}-catenin/Tcf$ reporter plasmid (P < 0.05). In conclusion, the lncRNA H19/miR-29b-3p/PGRN/Wnt axis counted a great deal for seeking appropriate diagnostic biomarkers and treatment targets for CRC.

• The Korean Journal of Emergency Medical Services
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• v.12 no.1
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• pp.17-26
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• 2008

Purpose : With a view to providing basic data to develop basic cardiopulmonary resuscitation education suitable for female college students, the basic cardiopulmonary resuscitation education was conducted to grasp students' knowledge, skill accuracy and the attitude change before and after the education. Methods : Convenience sampling was made on first graders(total-38 students) of K college located in K city, Chungcheongnam-do, and this was a preliminary research designed before and after choosing a single group. In terms of methods, the researchers as specified ; 1) Handed out questionnaires to students directly to make them fill in firsthand and collected the questionnaires. 2) Utilized Power Point materials based on 2005 AHA guideline and used Anne/Skill Reporter$^{(R)}$ torso produced by Leardal Inc, and Little Anne$^{(R)}$ to conduct practical education individually. 3) Asked students to give Anne/Skill Reporter$^{(R)}$ torso basic cardiopulmonary resuscitation five times with the ratio of 30:2, and then one of researchers filled in the evaluation sheet individually. 4) Evaluated the accuracy of students' ability to perform the resuscitation based on the record of Anne/Skill Reporter$^{(R)}$ integrated printer(which was the objective tool to grasp students' skills accuracy). 5) Gave out questionnaires to make students fill them in and then collected them, after completing the practical evaluation. Results : 1) In case of the attitude(confidence) about basic cardiopulmonary resuscitation, students' confidency rose from 14%(before the education) to 55.5%(after the education)- which was a positive change. 2) In case of the attitude(educational necessity) about basic cardiopulmonary resuscitation, students' confidency rose from 94.7%(before the education) to 100%(after the education)- which was a positive change. 3) As a result of the education, some female college students' scored knowledge about basic cardiopulmonary resuscitation. The average point also reached 88.95%(after the education), jump from 63.51%(before the education), which was the rise of 25.44%. 4) Regarding skill accuracy, pressure accuracy(%)($M{\pm}SD=91.37{\pm}14.16$) was higher than respiration accuracy(%)($M{\pm}SD=61.55{\pm}26.13$). Conclusion : The result showed that students' attitude(confidence, on basic cardiopulmonary resuscitation changed positively, and meaningful difference(p= .000) existed in the change of students' knowledge. Anne/Skill Reporter$^{(R)}$ performance showed that the accuracy of mouth-to-mouth resuscitation was lower than that of chest compression.

• Korean Journal of Radiology
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• v.21 no.6
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• pp.726-735
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• 2020

Objective: Recent innovations in biology are boosting gene and cell therapy, but monitoring the response to these treatments is difficult. The purpose of this study was to find an MRI-reporter gene that can be used to monitor gene or cell therapy and that can be delivered without a viral vector, as viral vector delivery methods can result in long-term complications. Materials and Methods: CMV promoter-human organic anion transporting polypeptide 1B3 (CMV-hOATP1B3) cDNA or CMV-blank DNA (control) was transfected into HEK293 cells using Lipofectamine. OATP1B3 expression was confirmed by western blotting and confocal microscopy. In vitro cell phantoms were made using transfected HEK293 cells cultured in various concentrations of gadoxetic acid for 24 hours, and images of the phantoms were made with a 9.4T micro-MRI. In vivo xenograft tumors were made by implanting HEK293 cells transfected with CMV-hOATP1B3 (n = 4) or CMV-blank (n = 4) in 8-week-old male nude mice, and MRI was performed before and after intravenous injection of gadoxetic acid (1.2 µL/g). Results: Western blot and confocal microscopy after immunofluorescence staining revealed that only CMV-hOATP1B3-transfected HEK293 cells produced abundant OATP1B3, which localized at the cell membrane. OATP1B3 expression levels remained high through the 25th subculture cycle, but decreased substantially by the 50th subculture cycle. MRI of cell phantoms showed that only the CMV-hOATP1B3-transfected cells produced a significant contrast enhancement effect. In vivo MRI of xenograft tumors revealed that only CMV-hOATP1B3-transfected HEK293 tumors demonstrated a T1 contrast effect, which lasted for at least 5 hours. Conclusion: The human endogenous OATP1B3 gene can be non-virally delivered into cells to induce transient OATP1B3 expression, leading to gadoxetic acid-mediated enhancement on MRI. These results indicate that hOATP1B3 can serve as an MRI-reporter gene while minimizing the risk of long-term complications.