• Title/Summary/Keyword: KB cells

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Rapid diagnosis of bovine tuberculosis in slaughter cattle using PCR (PCR 기법을 이용한 도축 소의 결핵병 신속진단)

  • Koh, Ba-Ra-Da;Kim, Hyun-Joong;Park, Duk-Woong;Park, Seong-Do;Kim, Jae-Ik;Park, Jong-Tae;Kim, Yong-Hwan
    • Korean Journal of Veterinary Service
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    • v.30 no.3
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    • pp.393-406
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    • 2007
  • Bovine tuberculosis is an important zoonosis worldwide. Mycobacterium bovis, the causative agent of this disease in cattle, is also a pathogen for humans and several economically important animals. The cases of tuberculosis are reported in two cow found at slaughter house located in Gwangju city. Histopathologically, in the lymph nodes, granulomas consisted of large areas of necrosis surrounded by variable thick bands of cellular infiltrate containing macrophages, Langhans-type multinucleated giant cells and lymphocytes. Lesions in the lung followed the same developmental pattern as did lesions in the lymph nodes with some exceptions. With the acid-fast staining, numerous mycobacteria were revealed in the lung and lymph nodes. M bovis was confirmed as a causative agent in these cattle using bacterial isolation and PCR and restriction fragment length polymorphism method based on a unique 12.7 kb fragment insertion sequence from the Mycobacterium tuberculosis genome and the pncA polymorphism, The insertion element IS6110 and IS1081 were present M bovis isolated from lungs and lymph nodes of cattle using PCR assay. These cases are interesting and important in public health aspect that M bovis-infected cattle were found during a routine post-mortem inspection at slaughter house.

Antimicrobial Effects of Oleanolic Acid against Streptococcus mutans and Streptococcus sobrinus Isolated from a Korean Population

  • Kim, Min-Jung;Kim, Chun-Sung;Ha, Woo-Hyung;Kim, Byung-Hoon;Lim, Yun-Kyong;Park, Soon-Nang;Cho, Yu-Jin;Kim, Myung-Mi;Ko, Jang-Hyuk;Kwon, Soon-Sung;Ko, Yeong-Mu;Kook, Joong-Ki
    • International Journal of Oral Biology
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    • v.35 no.4
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    • pp.191-195
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    • 2010
  • Oleanolic acid is a natural triterpenoid that exists widely in foods and some medicinal herbs. The purpose of this study was to determine the antimicrobial activity of oleanolic acid against Streptococcus mutans strains isolated from a Korean population. Antimicrobial activity against these bacteria was evaluated by minimal inhibitory concentration (MIC) and time kill curves. The tolerance of human gingival fibroblasts and human periodontal ligaments to oleanolic acid was tested using a methyl thiazolyl tetrazolium (MTT) assay. The $MIC_{90}$ value of oleanolic acid for both S. mutans and S. sobrinus isolated from Koreans was 8 ${\mu}g/ml$. Oleanolic acid showed bactericidal effects against S. mutans ATCC $25175^T$ and S. sobrinus ATCC $33478^T$ at $1\;{\times}\;MIC$ ($8{\mu}g/ml$) and had no cytotoxic effects against KB cells at this dose. The results suggest that oleanolic acid could be useful in the future development of oral hygiene products for the prevention of dental caries.

Crystal Structure and Comparative Sequence Analysis of GmhA from Colwellia psychrerythraea Strain 34H Provides Insight into Functional Similarity with DiaA

  • Do, Hackwon;Yun, Ji-Sook;Lee, Chang Woo;Choi, Young Jun;Kim, Hye-Yeon;Kim, Youn-Jung;Park, Hyun;Chang, Jeong Ho;Lee, Jun Hyuck
    • Molecules and Cells
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    • v.38 no.12
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    • pp.1086-1095
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    • 2015
  • The psychrophilic organism Colwellia psychrerythraea strain 34H produces extracellular polysaccharide substances to tolerate cold environments. Sedoheptulose 7-phosphate isomerase (GmhA) is essential for producing $\small{D}$-glycero-$\small{D}$-mannoheptose 7-phosphate, a key mediator in the lipopolysaccharide biosynthetic pathway. We determined the crystal structure of GmhA from C. psychrerythraea strain 34H (CpsGmhA, UniProtKB code: Q47VU0) at a resolution of $2.8{\AA}$. The tetrameric structure is similar to that of homologous GmhA structures. Interestingly, one of the catalytic residues, glutamate, which has been reported to be critical for the activity of other homologous GmhA enzymes, is replaced by a glutamine residue in the CpsGmhA protein. We also found differences in the conformations of several other catalytic residues. Extensive structural and sequence analyses reveal that CpsGmhA shows high similarity to Escherichia coli DnaA initiatorassociating protein A (DiaA). Therefore, the CpsGmhA structure reported here may provide insight into the structural and functional correlations between GmhA and DiaA among specific microorganisms.

Chloroplast Genome Evolution in Early Diverged Leptosporangiate Ferns

  • Kim, Hyoung Tae;Chung, Myong Gi;Kim, Ki-Joong
    • Molecules and Cells
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    • v.37 no.5
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    • pp.372-382
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    • 2014
  • In this study, the chloroplast (cp) genome sequences from three early diverged leptosporangiate ferns were completed and analyzed in order to understand the evolution of the genome of the fern lineages. The complete cp genome sequence of Osmunda cinnamomea (Osmundales) was 142,812 base pairs (bp). The cp genome structure was similar to that of eusporangiate ferns. The gene/intron losses that frequently occurred in the cp genome of leptosporangiate ferns were not found in the cp genome of O. cinnamomea. In addition, putative RNA editing sites in the cp genome were rare in O. cinnamomea, even though the sites were frequently predicted to be present in leptosporangiate ferns. The complete cp genome sequence of Diplopterygium glaucum (Gleicheniales) was 151,007 bp and has a 9.7 kb inversion between the trnL-CAA and trnV-GCA genes when compared to O. cinnamomea. Several repeated sequences were detected around the inversion break points. The complete cp genome sequence of Lygodium japonicum (Schizaeales) was 157,142 bp and a deletion of the rpoC1 intron was detected. This intron loss was shared by all of the studied species of the genus Lygodium. The GC contents and the effective numbers of codons (ENCs) in ferns varied significantly when compared to seed plants. The ENC values of the early diverged leptosporangiate ferns showed intermediate levels between eusporangiate and core leptosporangiate ferns. However, our phylogenetic tree based on all of the cp gene sequences clearly indicated that the cp genome similarity between O. cinnamomea (Osmundales) and eusporangiate ferns are symplesiomorphies, rather than synapomorphies. Therefore, our data is in agreement with the view that Osmundales is a distinct early diverged lineage in the leptosporangiate ferns.

Characterization of Melon necrotic spot virus Occurring on Watermelon in Korea

  • Kwak, Hae-Ryun;Kim, Jeong-Soo;Cho, Jeom-Deog;Lee, Joong-Hwan;Kim, Tae-sung;Kim, Mi-Kyeong;Choi, Hong-Soo
    • The Plant Pathology Journal
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    • v.31 no.4
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    • pp.379-387
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    • 2015
  • Melon necrotic spot virus (MNSV) was recently identified on watermelon (Citrullus vulgaris) in Korea, displaying as large necrotic spots and vein necrosis on the leaves and stems. The average occurrence of MNSV on watermelon was found to be 30-65% in Hapcheon and Andong City, respectively. Four isolates of the virus (MNSV-HW, MNSV-AW, MNSV-YW, and MNSV-SW) obtained from watermelon plants in different areas were non-pathogenic on ten general indicator plants, including Chenopodium quinoa, while they infected systemically six varieties of Cucurbitaceae. The virus particles purified by 10-40% sucrose density gradient centrifugation had a typical ultraviolet spectrum, with a minimum at 245 nm and a maximum at 260 nm. The morphology of the virus was spherical with a diameter of 28-30 nm. Virus particles were observed scattered throughout the cytoplasm of watermelon cells, but no crystals were detected. An ELISA was conducted using antiserum against MNSV-HW; the optimum concentrations of IgG and conjugated IgG for the assay were $1{\mu}l/ml$ and a 1:8,000-1:10,000 dilutions, respectively. Antiserum against MNSV-HW could capture specifically both MNSV-MN from melon and MNSV-HW from watermelon by IC/RT-PCR, and they were effectively detected with the same specific primer to produce product of 1,172 bp. The dsRNA of MNSV-HW had the same profile (4.5, 1.8, and 1.6 kb) as that of MNSV-MN from melon. The nucleotide sequence of the coat protein of MNSV-HW gave a different phylogenetic tree, having 17.2% difference in nucleotide sequence compared with MNSV isolates from melon.

Suppression of Protein Kinase C and Nuclear Oncogene Expression as Possible Action Mechanisms of Cancer Chemoprevention by Curcumin

  • Lin, Jen-Kun
    • Archives of Pharmacal Research
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    • v.27 no.7
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    • pp.683-692
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    • 2004
  • Curcumin (diferuloylmethane) is a major naturally-occurring polyphenol of Curcuma species, which is commonly used as a yellow coloring and flavoring agent in foods. Curcumin has shown anti-carcinogenic activity in animal models. Curcumin possesses anti-inflammatory activity and is a potent inhibitor of reactive oxygen-generating enzymes such as lipoxygenase/cyclooxygenase, xanthine dehydrogenase/oxidase and inducible nitric oxide synthase; and an effective inducer of heme oxygenase-1. Curcumin is also a potent inhibitor of protein kinase C(PKC), EGF(Epidermal growth factor)-receptor tyrosine kinase and LĸB kinase. Subsequently, curcumin inhibits the activation of NF(nucleor factor)KB and the expressions of oncogenes including c-jun, c-fos, c-myc, NIK, MAPKs, ERK, ELK, PI3K, Akt, CDKs and iNOS. It is proposed that curcumin may suppress tumor promotion through blocking signal transduction path-ways in the target cells. The oxidant tumor promoter TPA activates PKC by reacting with zinc thiolates present within the regulatory domain, while the oxidized form of cancer chemopreventive agent such as curcumin can inactivate PKC by oxidizing the vicinal thiols present within the catalytic domain. Recent studies indicated that proteasome-mediated degradation of cell proteins playa pivotal role in the regulation of several basic cellular processes including differentiation, proliferation, cell cycling, and apoptosis. It has been demonstrated that curcumin-induced apoptosis is mediated through the impairment of ubiquitin-proteasome pathway. Curcumin was first biotransformed to dihydrocurcumin and tetrahydrocurcumin and that these compounds subsequently were converted to monoglucuronide conjugates. These results suggest that curcumin-glucuronide, dihydrocurcumin-glucuronide, tetrahydrocurcumin-glucuronide and tetrahydrocurcumin are the major metabolites of curcumin in mice, rats and humans.

Cloning and Characterization of Porcine Uroplakin II Gene

  • D. N. Kwon;H. K. Shin;C. K. Hwang;D. W. Ok;Kim, J. H.
    • Proceedings of the KSAR Conference
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    • 2001.03a
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    • pp.19-19
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    • 2001
  • Mammalian urothelium undergoes unique membrane specialization by making the asymmetric unit membrane (AUM) that is covered with the apical cell surface during terminal differentiation. The AUM contains several major integral membrane proteins including uroplakin Ia, Ib, II and III. The genes for uroplakins have been cloned from humans and mice, but not from porcine. In this study, we report the cloning of the UPII genomic DNA, which codes for the full length open reading frame for the uroplakin II protein. The deduced amino acid sequence encodes of a hydrophobic NH$_2$-terminal peptide, a prosequence, and a mature protein. The prosequence contains three potential N-glycosylation sites and a RGRR cleavage site that may be involved in uroplakin II processing and maturation. Northern and immunohistochemistry analyses showed that the porcine UPII gene is only expressed in urothelium and that the protein was specifically localized in urothelial superficial cells. A 2kb of upstream in the promoter sequence contains multiple transcription factor binding sites, including GC-box, SPI, AP2, and GATA-box sites, but not for TATA or CAAT-box sequences. Comparison of the porcine UPII promoter sequence with that of the murine by MEME system presented two conserved motifs, suggesting a cis-acting regulatory role for the conserved sequences. Sequence homology between two species in motif A and B was 79% and 80% respectively, although their relative locations were different. During the gestation, mouse bladder at estrus stages and day 10 after parturition showed higher UPII expression, while showed lower expression at peri-implantation stage. Taken together, our results showed that the porcine UPII gene was expressed highly and specifically in the bladder urothelium and that steroid hormones for implantation changed the expression of UPII in the bladder, although the biological significance of UPII remains to be not determined.

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Characterization of Purple-discolored, Uppermost Leaves of Soybean; QTL Mapping, HyperspectraI Imaging, and TEM Observation

  • JaeJin Lee;Jeongsun Lee;Seongha Kwon;Heejin You;Sungwoo Lee
    • Proceedings of the Korean Society of Crop Science Conference
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    • 2022.10a
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    • pp.187-187
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    • 2022
  • Purple-discoloration of the uppermost leaves has been observed in some soybean cultivars in recent years. The purpose of this study was to characterize the novel phenotypic changes between the uppermost and middle leaves via multiple approaches. First, quantitative trait loci mapping was conducted to detect loci associated with the novel phenotype using 85 recombinant inbred lines (RILs) of the 'Daepung' × PI 96983 population. 180K SNP data, a major quantitative trait locus (QTL) was identified at around 60 cM of chromosome 6, which accounts for 56% of total phenotypic variance. The genomic interval is about ~700kb, and a list of annotated genes includes the T-gene which is known to control pubescence and seed coat color and is presumed to encode flavonoid 35-hydroxylase (F3'H). Based on Hyperspectral imaging, the reflectance at 528-554 nm wavelength band was extremely reduced in the uppermost leaves compared to the middle (green leaves), which is presumed die to the accumulation of anthocyanins. In addition, purple-discolored leaf tissues were observed and compared to normal leaves using a transmission electronic microscope (TEM). Base on observations of the cell organelles, the purple-discolored uppermost leaves had many pigments formed in the epidermal cells unlike the normal middle leaves, and the cell wall thickness was twice as thick in the discolored leaves. The thickness of the thylakoid layer in the chloroplast the number of starch grains, the size of starch all decreased in the discolored leaves, while the number of plastoglobule and mitochondria increased.

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Extraction & Purification of ${\beta}$-carotene from Recombinant Escherichia coli (재조합 대장균으로부터 고순도 베타-카로틴의 추출 및 정제)

  • Jo, Ji-Song;Nguyen, Do Quynh Anh;Yun, Jun-Ki;Kim, Yu-Na;Kim, You-Geun;Kim, Sung-Bae;Seo, Yang-Gon;Lee, Byung-Hak;Kang, Moon-Kook;Kim, Chang-Joon
    • Microbiology and Biotechnology Letters
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    • v.37 no.3
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    • pp.231-237
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    • 2009
  • This paper aimed to develop a solvent extraction and purification process to recover high-purified ${\beta}$-carotene from recombinant Escherichia coli. Cells harvested from the culture broth were treated through numerous steps: dehydration, solvent extraction, crystal formation and separation. To optimize the extracting condition, experiments were carried out to investigate the effect of cell disruption, temperature, organic solvents, solvent-biomass ratio on the yield of ${\beta}$-carotene extracted from cells. The result indicated that no significant differences of extraction yield were observed from cells with or without step of cell disruption. Among different extracting solvents, the highest extraction yield of ${\beta}$-carotene, 30.3 mg-${\beta}$-carotene/g-dry cells, was obtained with isobutyl acetate at solvent-biomass ratio 25 mL/g-dry cells at $50^{\circ}C$. Notably, in case of acetone, the extraction yield was quite low when using acetone itself, but increased almost up to the highest value when combining this solvent and olive oil. The purity of ${\beta}$-carotene crystals obtained from crystallization and separation was 89%. The purity degree was further improved up to 98.5% by treating crude crystals with additional ethanol washing.

NF-${\kappa}B$ Activation and cIAP Expression in Radiation-induced Cell Death of A549 Lung Cancer Cells (A549 폐암세포주의 방사선-유도성 세포사에서 NF-${\kappa}B$ 활성화 및 cIAP 발현)

  • Lee, Kye Young;Kwak, Shang-June
    • Tuberculosis and Respiratory Diseases
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    • v.55 no.5
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    • pp.488-498
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    • 2003
  • Background : Activation of the transcription factor NF-${\kappa}B$ has been shown to protect cells from tumor necrosis factor-alpha, chemotherapy, and radiation-induced apoptosis. NF-${\kappa}B$-dependent cIAP expression is a major antiapoptotic mechanism for that. NF-${\kappa}B$ activation and cIAP expression in A549 lung cancer cells which is relatively resistant to radiation-induced cell death were investigated for the mechanism of radioresistance. Materials and methods : We used A549 lung cancer cells and Clinac 1800C linear accelerator for radiation. Cell viability test was done by MTT assay. NF-${\kappa}B$ activation was tested by luciferase reporter gene assay, Western blot for $I{\kappa}B{\alpha}$ degradation, and electromobility shift assay. For blocking ${\kappa}B$, MG132 and transfection of $I{\kappa}B{\alpha}$-superrepressor plasmid construct were used. cIAP expression was analyzed by RT-PCR and cIAP2 promoter activity was performed using luciferase assay system. Results : MTT assay showed that cytotoxicity even 48 hr after radiation in A549 cells were less than 20%. Luciferas assay demonstrated weak NF-${\kappa}B$ activation of $1.6{\pm}0.2$ fold compared to PMA-induced $3.4{\pm}0.9$ fold. Radiation-induced $I{\kappa}B{\alpha}$ degradation was observed in Western blot and NF-${\kappa}B$ DNA binding was confirmed by EMSA. However, blocking NF-${\kappa}B$ using MG132 and $I{\kappa}B{\alpha}$-superrepressor transfection did not show any sensitizing effect for radiation-induced cell death. The result of RT-PCR for cIAP1 & 2 expression was negative induction while TNF-${\alpha}$ showed strong expression for cIAP1 & 2. The cIAP2 promoter activity also did not show any change compared to positive control with TNF-${\alpha}$. Conclusion : We conclude that activation of NF-${\kappa}B$ does not determine the intrinsic radiosensitivity of cancer cells, at least for the cell lines tested in this study.