• Journal of Life Science
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• v.27 no.8
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• pp.857-863
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• 2017

The FBJ murine osteosarcoma viral oncogene homolog B (FosB) gene is located at chromosome 19, and encodes 43 Kda protein. Functionally, the FosB gene is important for differentiation, development, and pathogenesis. Furthermore, the FosB gene is suggested as possible biomarker for tracing disease prognosis. In this study, we constructed plasmid containing a FosB promoter region and evaluate its promoter activity. We analyzed the putative promoter region in FosB genomic DNA using bioinformatics program, and we found important regulatory elements in 1 Kb upstream from transcription start site (TSS). Therefore, we performed polymerase chain reaction (PCR) amplification on region from-1,555 upstream to +73 of the FosB genomic DNA, and PCR product was inserted into TA vector to create the $TA-1^{st}FosBp$ plasmid. We then prepared the primer sets, which contain a restriction enzyme site for Kpn1 and Nhe1, in order to reinsert into the TA vector to prepare $TA-2^{nd}FosBp$ plasmid. It was finally subcloned into pGL3-luc vector after enzyme cutting. To evaluate whether the cloned plasmid is useful in cell based experiment, we performed luciferase assay with pGL3-FosBp-luctransfection. FosB promoter activity was increased compared to empty vector, and this activity was significantly increased by treatment of doxorubicin and taxol. We obtained consistent data on regulation of FosB gene expression after anticancer drug treatment using Western blot analysis. The results suggest that promoter cloning of the human FosB gene is very useful for studying gene expression and analyzing biomarkers.

• Journal of fish pathology
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• v.10 no.2
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• pp.153-164
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• 1997

Characteristics and pathogenicity of Yersinia sp. strain isolated from diseased cultured olive flounder, Paralichthys olivaceous were studied. The causative organisms were identified as Yersinia sp. by biochemical and biophysical characteristics. The strain was named KF-1, and it showed the optimal growth rate at pH 9.0 and 1% NaCl. Total cell protein peptide bands of the Yersinia sp. KF-1 were between 14.4-100.6 Kd in molecular weight by the electrophoretic analysis, and a total of 28 bands appeared. The band found at 32.8 Kd in molecular weight was the major one of electrophoretic phase. In the pathogenicity test of the isolate to the flounder injecting with $1.0{\times}10^7$ cfu/fish 9 died out of 10 within 60 hrs, and in the group with $1.0{\times}10^7$ cfu/fish 5 died within 60 hrs. Thus the $LD_{50}$ was presumed to be $1.0{\times}10^7$ cfu/fish. In the drug sensitivity test KF-1 strain was sensitive to chloramphenicol, gentamycin, kanamycin, streptomycin and pefloxacin, and intermediate to erythromycin, and resistant to ampicillin, cephalothin, sulfamethoxazole/trimethoprim, tetracycline and vancomycin.

• Biomedical Science Letters
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• v.11 no.4
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• pp.465-471
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• 2005

Ofloxacin has antimycobacterial activity that possibly contributes a pivotal role in the second-line drug regimens that are used for the treatment of multidrug-resistant tuberculosis. However, in some communities, the resistance rate of Mycobacterium tuberculosis to this agent is surging. Therefore, a rapid and accurate method that can be used to determine the resistance of M tuberculosis to the ofloxacin can be very useful for effective treatment of the patients. As an effort to develop such a method, this study was set up to reveal general types of mutations that are related to ofloxacin resistance of M tuberculosis. From previous studies, it has been well known that ofloxacin resistance is associated with mutations in a gene encoding the gyrase A subunit protein. In this study, we obtained 43 ofloxacin-resistant and 50 ofloxacin-susceptible M tuberculosis clinical isolates from Masan National TB Hospital, and sequences of DNA fragment of 320 bp, region of gyrA corresponding to the ofloxacin resistance-determining region were analyzed. In brief, the results showed that a total of seven mutation types were found at gyrA. Theses mutations were all clustered within nucleotides 2574 to 2586 of the gyrA gene (codons 88 to 94). Codon 94 was the most frequently substituted site. Twenty-four of the 43 isolates had mutations at this position resulting in a total of five different types of amino acid changes $(Asp{\to}Ala,\;Asp{\to}Gly,\;Asp{\to}His,\;Asp{\to}Tyr,\;and\;Asp{\to}Asn)$. Five isolates contained a mutation at codon 90 resulting $Ala{\to}Val$ change. Four isolates had mutations at codon 91 causing a $Ser{\to}Pro$ change at this site. Two isolates contained a mutation at codon 88 and each of them resulted in different types of amino acid changes $(Gly{\to}Cys,\;Gly{\to}Ala)$. On the other hand, polymorphic site at codon 95 was found in both ofloxacin-resistant and ofloxacin-susceptible isolates. From these results, we concluded that the rate of mutations present in gyrA among ofloxacin-resistant M. tuberculosis in Korea is similar to the general rates of mutations found throughout the world. Subsequently, an oligonucleotide probe was designed based on the results of sequence analysis and was used to develop a dot blot hybridization assay system to determine ofloxacin-resistance of M tuberculosis. To evaluate this probe, dot-blot hybridization was carried out using other 57 clinical isolates, and the results showed that the dot-blot hybridization assay is good for detecting sequence alterations atgyrA gene.

• Biomedical Science Letters
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• v.17 no.4
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• pp.297-304
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• 2011

Influenza A virus of the Orthomyxoviridae family is a contagious respiratory pathogen that continues to evolve and burden in the human public health. It is able to spread efficiently from human to human and have the potential to cause pandemics with significant morbidity and mortality. It has been estimated that every year about 500 million people are infected with this virus, causing about approximately 0.25 to 0.5 million people deaths worldwide. Influenza A viruses are classified into different subtypes by antigenicity based on their hemagglutinin (HA) and neuraminidase (NA) proteins. The sudden emergence of influenza A virus subtypes and access for epidemiological analysis of this subtypes demanded a rapid development of specific diagnostic tools. Also, rapid identification of the subtypes can help to determine the antiviral treatment, because the different subtypes have a different antiviral drug resistance patterns. In this study, our aim is to detect influenza A virus subtypes by using real-time nucleic acid sequence based amplification (NASBA) which has high sensitivity and specificity through molecular beacon. Real-time NASBA is a method that able to shorten the time compare to other molecular diagnostic tools and is performed by isothermal condition. We selected major pandemic influenza A virus subtypes, H3N2 and H5N1. Three influenza A virus gene fragments such as HA, NA and matrix protein (M) gene were targeted. M gene is distinguished influenza A virus from other influenza virus. We designed specific primers and molecular beacons for HA, NA and M gene, respectively. In brief, the results showed that the specificity of the real-time NASBA was higher than reverse transcription polymerase chain reaction (RT-PCR). In addition, time to positivity (TTP) of this method was shorter than real-time PCR. This study suggests that the rapid detection of neo-appearance pandemic influenza A virus using real-time NASBA has the potential to determine the subtypes.

• Journal of Society of Preventive Korean Medicine
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• v.11 no.2
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• pp.23-39
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• 2007

Objectives : The aim of this study was to evaluate effects of herbal medicine and other associated factors for abnormal liver function tests(especially total bilirubin, AST, ALT, and ${\gamma}GT$) levels in outpatients at an oriental medical clinic. Methods : A cross-sectional study based on clinical records was conducted on 1,871 patients at one Oriental medical clinic in Seoul, Korea. 504 patients received a liver function tests for screening and 497 patients ranging from the age of 4 to 74 were chosen for the study. Patients with basal liver disease or DM were excluded during the screening process. Patients were classified into case(abnormal) and control(normal) groups by normal liver function test references. Results and Conclusions : In this study, 33.0% of the patients were females, and 67.0% were males. The mean age was $34.7{\pm}11.9$ years old. The mean total protein value was $6.78{\pm}0.66g/dL$(male $6.79{\pm}0.61$, female $6.76{\pm}0.76$), albumin $3.89{\pm}0.50g/dL$(male $3.94{\pm}0.47$, female $3.81{\pm}0.54$), total bilirubin $0.51{\pm}0.35mg/dL$(male $0.54{\pm}0.39$, female $0.45{\pm}0.23$), AST $23.31{\pm}18.22U/L$(male $26.37{\pm}20.73$, female $17.09{\pm}8.72$), ALT $33.49{\pm}36.36U/L$(male $40.56{\pm}41.77$, female $19.13{\pm}12.64$), LDH $258.07{\pm}74.84mg/dL$(male $263.68{\pm}73.77$, female $246.70{\pm}75.92$), and ${\gamma}GT$ $39.64{\pm}59.16U/L$(male $50.15{\pm}69.43$, female $17.83{\pm}8.36$). The percentage of abnormal total bilirubin levels(>1.0) in these patients was 2.6%, abnormal AST levels(>39m, >29f) 8.5%, ALT levels(>47m, >32f) 18.8%, and ${\gamma}GT$ (>50m, >40f) 19.9%.

• Biomolecules & Therapeutics
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• v.24 no.4
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• pp.402-409
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• 2016

It has been found that 4-isopropyl-2,6-bis(1-phenylethyl)phenol (KTH-13), a novel compound isolated from Cordyceps bassiana, is able to suppress tumor cell proliferation by inducing apoptosis. To mass-produce this compound, we established a total synthesis method. Using those conditions, we further synthesized various analogs with structural similarity to KTH-13. In this study, we aimed to test their anti-cancer activity by measuring anti-proliferative and pro-apoptotic activities. Of 8 compounds tested, 4-methyl-2,6-bis(1-phenylethyl)phenol (KTH-13-Me) exhibited the strongest anti-proliferative activity toward MDA-MB 231 cells. KTH-13-Me also similarly suppressed the survival of various cancer cell lines, including C6 glioma, HCT-15, and LoVo cells. Treatment of KTH-13-Me induced several apoptotic signs in C6 glioma cells, such as morphological changes, induction of apoptotic bodies, and nuclear fragmentation and chromatin condensation. Concordantly, early-apoptotic cells were also identified by staining with FITC-Annexin V/PI. Moreover, KTH-13-Me highly enhanced the activation of caspase-3 and caspase-9, and decreased the protein level of Bcl-2. In addition, the phosphorylation levels of Src and STAT3 were diminished in KTH-13-Me-treated C6 cells. Therefore, these results suggest that KTH-13-Me can be developed as a novel anti-cancer drug capable of blocking proliferation, inducing apoptosis, and blocking cell survival signaling in cancer cells.

• Korean Journal of Microbiology
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• v.47 no.3
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• pp.249-254
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• 2011

Mycophenolic acid blocking the synthesis of xanthosine monophosphate is a nonnucleoside inhibitor of inosine monophosphate dehydrogenase. Therefore mycopholoic acid is a drug currently used as immunosuppressive agent in transplantation of heart, kidney and liver. Mycophenolic acid has been industrially produced through fermentation process by fungus Penicillium brevi-compactum. In this study, the profile of mycophenolic acid fermentation was observed in 5L-jar fermentor to investigate the utilization of carbon and nitrogen sources and the production of mycophenolic acid. It was investigated that what kind of carbon sources was better to cell growth and mycophenolic acid production. Fructose was the best carbon source for mycophenolic acid fermentation, but it is the most expensive one. Thereafter molasses containing sucrose as the supply source of fructose was confirmed to be the best carbon source for the industrial production. Use of molasses increased the fermentation yield of mycophenolic acid more than two times higher than glucose. It was confirmed that urea was the best inorganic nitrogen source, which did not give rise to sudden drop of culture pH. Addition of urea increased the fermentation yield of mycophenolic acid about 3.6 times higher than addition of ammonium nitrate as control. Casein, peptone and casamino acid originated from milk protein increased the fermentation yield of mycophenolic acid about 3.4 times higher than control. Peptone and casamino acid, which are casein hydrolysates, increased cell growth considerably as well.

• Journal of Pharmaceutical Investigation
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• v.41 no.4
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• pp.255-262
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• 2011

Method using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS) was developed and validated for the determination of pregabalin in plasma samples. Acquisition was performed by monitoring the transitions: m/z 160.1${\rightarrow}$142.2 for pregabalin and m/z 423.2${\rightarrow}$207.1 for losartan (as an internal standard). After cold acetonitrileinduced protein precipitation of the plasma samples, separation was performed with C18 column by isocratic mobile phase consisted of 10 mM ammonium acetate and acetonitrile (15:85, v/v). Results were linear over the concentration ranged from 0.1 to $10{\mu}g$/mL and the correlation coefficients (r) were $\geq0.99$. Intra- and inter-day precisions were $\leq6.02$ and $\leq11.04%$, respectively, and intra- and inter-day accuracies were 96.60-101.09 and 98.10-102.60%, respectively. This validated method was successfully applied to a bioequivalence study of two formulations of pregabalin, Daewoong pregabalin capsule (Daewoong Pharm. Co., Ltd.) and Lyrica$^{(R)}$ capsule (Pfizer Korea Ltd.) in twenty eight healthy Korean volunteers. The subjects received a single oral dose of each formulation (150 mg as pregabalin) in a randomized $2{\times}2$ crossover study and plasma samples were obtained from each subject at predetermined time intervals. Then, the pharmacokinetic parameters ($AUC_{0-t}$, $C_{max}$ and $T_{max}$) were calculated and statistically analyzed to assess the differences between two formulations. The 90% confidence intervals for the log-transformed data were acceptable range of log 0.8-log 1.25 (e.g., log 1.0048-log 1.0692 for AUC0-t, log 0.9142-log 1.0421 for $C_{max}$). Thus, $AUC_{0-t}$ and $C_{max}$ met the criteria of the Korea Food and Drug Administration (KFDA) for bioequivalence test indicating that Daewoong pregabalin capsule was bioequivalent to Lyrica$^{(R)}$ capsule.

• Proceedings of the Korean Society of Applied Pharmacology
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• 1996.04a
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• pp.19-29
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• 1996

The neurological manifestations of AIDS include dementia, encountered even in the absence of opportunistic superinfection or malignancy. The AIDS Dementia Complex appears to be associated with several neuropathological abnormalities, including astrogliosis and neuronal injury or loss. How can HIV-1 result in neuronal damage if neurons themselves are only rarely, if ever, infected by the vitus\ulcorner In vitro experiments from several different laboratiories have lent support to the existence of HIV- and immune-related toxins. In one recently defined pathway to neuronal injury, HIV-infected macrophages/microglia as well as macrophages activated by HIV-1 envelope protein gp120 appear to secrete excitants/neurotoxins. These substances may include arachidonic acid, platelet-activating factor, free radicals (NO - and O$_2$), glutamate, quinolinate, cysteine, cytokines (TNF-${\alpha}$, IL1-B, IL-6), and as yet unidentified factors emanating from stimulated macrophages and possibly reactive astrocytes. A final common pathway for newonal suscepubility appears to be operative, similar to that observed in stroke, trauma, epilepsy, and several neurodegenerative diseases, including Huntington's disease, Parkinson's disease, and amyotrophic lateral sclerosis. This mechanism involves excessive activation of N-methyl-D-aspartate (NMDA) receptor-operated channels, with resultant excessive influx of Ca$\^$2+/ leading to neuronal damage, and thus offers hope for future pharmacological intervention. This chapter reviews two clinically-tolerated NMDA antagonists, memantine and nitroglycerin; (ⅰ) Memantine is an open-channel blocker of the NMDA-associated ion channel and a close congener of the anti-viral and anti-parkinsonian drug amantadine. Memantine blocks the effects of escalating levels of excitotoxins to a greater degree than lower (piysiological) levels of these excitatory amino acids, thus sparing to some extent normal neuronal function. (ⅱ) Niuoglycerin acts at a redox modulatory site of the NMDA receptor/complex to downregulate its activity. The neuroprotective action of nitroglycerin at this site is mediated by n chemical species related to nitric oxide, but in a higher oxidation state, resulting in transfer of an NO group to a critical cysteine on the NMDA receptor. Because of the clinical safety of these drugs, they have the potential for trials in humans. As the structural basis for redox modulation is further elucidated, it may become possible to design even better redox reactive reagents of chinical value. To this end, redox modulatory sites of NMDA receptors have begun to be characterized at a molecular level using site-directed mutagenesis of recombinant subunits (NMDAR1, NMDAR2A-D). Two types of redox modulation can be distinguished. The first type gives rise to a persistent change in the functional activity of the receptor, and we have identified two cysteine residues on the NMDARI subunit (#744 and #798) that are responsible for this action. A second site, presumably also a cysteine(s) because <1 mM N-ethylmaleimide can block its effect in native neurons, underlies the other, more transient redox action. It appears to be at this, as yet unidentified, site on the NMDA receptor that the NO group acts, at least in recombinant receptors.

• Herbal Formula Science
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• v.28 no.4
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• pp.339-349
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• 2020

Objectives : Wiryeong-tang (WRT) is a traditional herbal medicine used to treat kidney-related diseases. However, the anti-inflammatory and anti-gastritis effect of Wiryeong-tang was not well known. Therefore, we experimented to confirmed the anti-inflammatory and anti-gastritis effects of Wiryeong-tang. Methods : The RAW 264.7 cells were pre treated with Wiryeong-tang mix soft extract (WRT-mse; 50, 100 and 200 ㎍/mL) for 1 hrs, and then incubated with lipopolysaccharide (LPS; 500 ng/mL). Cell viability was measured by the MTT method, and nitric oxide (NO) was measured with griess reagent. In addition, pro-inflammatory cytokines were measured by enzyme-linked immunosorbent assay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR). For anti-gastritis effect in vivo, acute gastritis was induced using 150 mM HCl/60% ethanol used ICR mice. WRT-mse (133 mg/kg) was pre treated for 3 days and then treated with 150 mM HCl/60% ethanol 1 hrs later. Then gastritis was observed and inflammatory cytokines in the gastric tissue was measured. Results : The 8 marker components of the WRT-mse were determined by simultaneous analysis using HPLC. WRT-mse was not toxic and inhibited pro-inflammatory cytokines such as IL-1β, IL-6 and TNF-α at NO production, protein and mRNA levels. Also, it was confirmed that WRT-mse improved bleeding and edema in gastritis, and suppresses inflammatory cytokines. Conclusion : In summary, our results suggest that the treatment of the WRT-mse reduced and improved the 150 mM HCl/60% ethanol induced acute gastritis and the inflammation caused by LPS stimulation in RAW 264.7 cells. Therefore, this study may provide useful drug or clinical evidence for WRT-mse to prevent inflammation.