• Title/Summary/Keyword: Cell based assay

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Low Level of TERC Gene Amplification between Chronic Myeloid Leukaemia Patients Resistant and Respond to Imatinib Mesylate Treatment

  • Mohamad Ashari, Zaidatul Shakila;Sulong, Sarina;Hassan, Rosline;Husin, Azlan;Sim, Goh Ai;Wahid, S. Fadilah Abdul
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.4
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    • pp.1863-1869
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    • 2014
  • The amplification of telomerase component (TERC) gene could play an important role in generation and treatment of haematological malignancies. This present study was aimed to investigate copy number amplification status of TERC gene in chronic myeloid leukaemia (CML) patients who were being treated with imatinib mesylate (IM). Genomic DNA was extracted from peripheral blood of CML-IM Resistant (n=63), CML-IM Respond (n=63) and healthy individuals (n=30). TERC gene copy number predicted (CNP) and copy number calculated (CNC) were determined based on $Taqman^{(R)}$ Copy Number Assay. Fluorescence in situ hybridization (FISH) analysis was performed to confirm the normal signal pattern in C4 (calibrator) for TERC gene. Nine of CML patients showed TERC gene amplification (CNP=3), others had 2 CNP. A total of 17 CML patients expressed CNC>2.31 and the rest had 2.31>CNC>1.5. TERC gene CNP value in healthy individuals was 2 and their CNC value showed in range 1.59-2.31. The average CNC TERC gene copy number was 2.07, 1.99 and 1.94 in CML-IM Resistant patients, CML-IM Respond and healthy groups, respectively. No significant difference of TERC gene amplification observed between CML-IM Resistant and CML-IM Respond patients. Low levels of TERC gene amplification might not have a huge impact in haematological disorders especially in terms of resistance towards IM treatment.

The effect of nitrogen-fixing microorganisms on plant promotion in cabbage

  • Moon, Je-Hun;Jadamba, Chuluuntsetseg;Yoo, Soo-Cheul
    • Proceedings of the Korean Society of Crop Science Conference
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    • 2017.06a
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    • pp.190-190
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    • 2017
  • Chemical fertilizers have been used to increase crop production and contributed to escaping food shortages. However, excessive use of chemical fertilizers over a long period caused many problems such as environmental pollution and the hampered production potential of the land. Thus, it is necessary to develop eco-friendly bio-fertilizers that can replace the use of chemical fertilizers. Here, we tested the effect of some nitrogen-fixing microorganims on the plant growth promotion. Seventy free-living nitrogen fixing microorganisms were isolated from rhizosphere of crop cultivation fields, streamside soils and sludge in Ansung, Korea. Of them, three strains (NF2-4-1, Yeast; EMM409, Mesorhizobium; Gsoil662, Burkholderia) were selected to be most efficient in the capacity of N-fixing nitrogen based on colony forming cell assay in N-free media. To investigate the ability to promote plant growth, these strains were inoculated into the soil and cabbage were grown for 4 weeks in the grown chamber. Fresh weight, dry weight, and leaf area were measured from 4-week-old plants. Phenotypic analysis revealed that the growth of the plants inoculated with NF2-4-1 and EMM409 strains were significantly promoted compared to the mock-treated control plants, while Gsoil662-inoculated plants did not show statically significant promotion. These results indicate that these nitrogen-fixing microorganims can be used to develop plant growth promoting bio-fertilizers. Further analysis on nitrogen fixing level in soil by these strains will be tested.

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Cholera Toxin B Subunit-Porphyromonas gingivalis Fimbrial Antigen Fusion Protein Production in Transgenic Potato

  • Lee, Jin-Yong;Kim, Mi-Young;Jeong, Dong-Keun;Yang, Moon-Sik;Kim, Tae-Geum
    • Journal of Plant Biotechnology
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    • v.36 no.3
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    • pp.268-274
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    • 2009
  • Porphyromonas gingivalis, the gram-negative anaerobic oral bacterium, initiates periodontal disease by binding to saliva-coated oral surface. The cholera toxin B subunit (CTB) genetically linked to FimA1 (1-200 aa) or FimA2 (201-337 aa) of the P. gingivalis fimbrial antigen were introduced into Solanum tuberosum cells by Agrobacterium tumefaciens-mediated transformation method. The integration of CTB-FimA1 or CTB-FimA2 fusion genes were confirmed in the chromosome of transformed leaves by genomic DNA PCR amplification method. Synthesis and assembly of the CTB-FimA fusion proteins into oligomeric structures with pentamer size was detected in transformed tuber extracts by immunoblot analysis. The binding activities of CTB-FimA fusion proteins to intestinal epithelial cell membrane receptors were confirmed by GM1-ganglioside enzyme-linked immunosorbent assay (GM1-ELISA). The ELISA showed that the expression levels of the CTB-FimA1 or CTB-FimA2 fusion proteins were 0.0019, 0.002% of the total soluble protein in transgenic tuber tissues, respectively The synthesis of CTB-FimA monomers and their assembly into biologically active oligomers in transformed potato tuber tissues demonstrates the feasibility of using edible plants for the production of enterocyte targeted fimbrial antigens that could elicit mucosal immune responses.

Investigation on Hydration Process and Biocompatibility of Calcium Silicate-Based Experimental Portland Cements

  • Lim, Jiwon;Guk, Jae-Geun;Singh, Bhupendra;Hwang, Yun-Chan;Song, Sun-Ju;Kim, Ho-Sung
    • Journal of the Korean Ceramic Society
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    • v.56 no.4
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    • pp.403-411
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    • 2019
  • In this work, the hydration process and cytotoxicity of lab-synthesized experimental Portland cements (EPCs) were investigated for dental applications. For this purpose, EPCs were prepared using laboratory-synthesized clinker constituents, tricalcium silicate (C3S), dicalcium silicate (C2S), and tricalcium aluminate (C3A). C-A was prepared by the Pechini method, whereas C3S and C2S were synthesized by solid-state reactions. The phase compositions were characterized by X-ray diffraction (XRD) analysis, and the hydration process of the individual constituents and their combinations, with and without the addition of gypsum, was investigated by electrochemical impedance spectroscopy (EIS). Furthermore, four EPC compositions were prepared using the lab-synthesized C-A, C3S, and C2S, and their hydration processes were examined by EIS, and their cytotoxicity to HPC and HIPC cells were tested by performing an XTT assay. None of the EPCs exhibited any significant cytotoxicity for 7 days, and no significant difference was observed in the cell viabilities of ProRoot MTA and EPCs. The results indicated that all the EPCs are sufficiently biocompatible with human dental pulp cells and can be potential substitutes for commercial dental cements.

Prevalence of fimA Genotypes of Porphyromonas gingivalis Strains in peri-implantitis patients (임플란트주위염시 Porphyromonas gingivalis 섬모유전형의 출현율)

  • Shin, Seung-Il;Kwon, Young-Hyuk;Park, Joon-Bong;Herr, Yeek;Chung, Jong-Hyuk
    • Journal of Periodontal and Implant Science
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    • v.35 no.1
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    • pp.31-41
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    • 2005
  • Fimbriae (fimA) of Porphyromonas gingivalis are filamentous components on the cell surface and are thought to play an important role in the colonization and invasion of periodontal tissue. P. gnigivalis fimA gene encoding fimbrillin, a subunit of fimbriae, has been classified into 5 genotypes (types I to V) based on the nucleotide sequences. In the present study, we examined the prevalence of these fimA genotypes in patients with dental implant and the relationship between prevalence of these genotypes and peri-implantitis. Dental plaque specimens obtained from 80 peri-implant sulci of 50 patients with dental implants were analyzed by 16S rRNA fimA gene-directed PCR assay. P. gingivalis were detected in 74.4% of the samples of the control group (healthy peri- implant sulci; probing depth<5mm) and in 92.0% of the samples of the test group (peri-implant sulci with peri-iimplantitis; probing $depth{\geqq}5mm$). Among the P. gingivalis-positive samples of the control group, the most prevalent fimA type was type I (29.3%), followed by type II (26.8%). In contrast, a majority among the P. gingivalis-positive samples of the test group was type II (56.S%), followed by type I (43.5%). TypeII fimA genotype organisms were detected more frequently in the test group and a significant difference in the occurrence of type II was observed between test and the control groups. A correlation between specific fimA types and peri-implant health status was found in type II (OR 3.545) and only a weak relationship was revealed in typeIV(OR 3.807). These findings indicate that P. gingivalis strains that possess type II fimA are predominant in peri-implant sulci with peri-implantitis and are closely associated with peri-implant health status. P. gingivalis with type II fimA may be involved in peri-implantitis.

Anti-aging Potential of Extracts Prepared from Fruits and Medicinal Herbs Cultivated in the Gyeongnam Area of Korea

  • Shon, Myung-Soo;Lee, Yunjeong;Song, Ji-Hye;Park, Taehyun;Lee, Jun Kyoung;Kim, Minju;Park, Eunju;Kim, Gyo-Nam
    • Preventive Nutrition and Food Science
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    • v.19 no.3
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    • pp.178-186
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    • 2014
  • Many recent studies have focused on maintaining a healthy life by preventing and/or postponing the aging process. Numerous studies have reported that continuous exposure to reactive oxygen species can stimulate skin aging and that excessive accumulation of fat can cause an impaired skin barrier and tissue structure alterations. Thus, the maintenance of antioxidant homeostasis and the suppression of adipose accumulation are important strategies for skin anti-aging. Here, we prepared three types of extracts [whole juice, acetone-perchloric acid (PCA), and ethanol] from 20 fruits and medicinal herbs native to the Gyeongnam area of Korea. The total phenolic content of each extract was analyzed, and we observed higher total phenolic contents in the medicinal herbs. Consistent with this, the results of the oxygen radical absorbance activity capacity assay indicated that the in vitro antioxidant activities of the medicinal herb extracts were stronger than those of the fruit extracts. The fruits and medicinal herbs had strong effects on cell-based systems, including $H_2O_2$-induced oxidative stress in human keratinocytes and 3T3-L1 lipid accumulation. Nishimura Wase persimmon, Taishu persimmon, wrinkled giant hyssop, sweet wormwood, Chinese cedar, red perilla, tan shen, hiyodori-jogo, and cramp bark may be natural anti-aging materials with effective antioxidant and anti-adipogenic activities. Taken together, our findings may provide scientific evidence supporting the development of functional foods and nutraceuticals from fruits and medicinal herbs.

Identification and Antimicrobial Activity Detection of Lactic Acid Bacteria Isolated from Corn Stover Silage

  • Li, Dongxia;Ni, Kuikui;Pang, Huili;Wang, Yanping;Cai, Yimin;Jin, Qingsheng
    • Asian-Australasian Journal of Animal Sciences
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    • v.28 no.5
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    • pp.620-631
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    • 2015
  • A total of 59 lactic acid bacteria (LAB) strains were isolated from corn stover silage. According to phenotypic and chemotaxonomic characteristics, 16S ribosomal DNA (rDNA) sequences and recA gene polymerase chain reaction amplification, these LAB isolates were identified as five species: Lactobacillus (L.) plantarum subsp. plantarum, Pediococcus pentosaceus, Enterococcus mundtii, Weissella cibaria and Leuconostoc pseudomesenteroides, respectively. Those strains were also screened for antimicrobial activity using a dual-culture agar plate assay. Based on excluding the effects of organic acids and hydrogen peroxide, two L. plantarum subsp. plantarum strains ZZU 203 and 204, which strongly inhibited Salmonella enterica ATCC $43971^T$, Micrococcus luteus ATCC $4698^T$ and Escherichia coli ATCC $11775^T$ were selected for further research on sensitivity of the antimicrobial substance to heat, pH and protease. Cell-free culture supernatants of the two strains exhibited strong heat stability (60 min at $100^{\circ}C$), but the antimicrobial activity was eliminated after treatment at $121^{\circ}C$ for 15 min. The antimicrobial substance remained active under acidic condition (pH 2.0 to 6.0), but became inactive under neutral and alkaline condition (pH 7.0 to 9.0). In addition, the antimicrobial activities of these two strains decreased remarkably after digestion by protease K. These results preliminarily suggest that the desirable antimicrobial activity of strains ZZU 203 and 204 is the result of the production of a bacteriocin-like substance, and these two strains with antimicrobial activity could be used as silage additives to inhibit proliferation of unwanted microorganism during ensiling and preserve nutrients of silage. The nature of the antimicrobial substances is being investigated in our laboratory.

Development and Characterization of a Novel Anti-idiotypic Monoclonal Antibody to Growth Hormone, Which Can Mimic Physiological Functions of Growth Hormone in Primary Porcine Hepatocytes

  • Lan, Hai-Nan;Jiang, Hai-Long;Li, Wei;Wu, Tian-Cheng;Hong, Pan;Li, Yu Meng;Zhang, Hui;Cui, Huan-Zhong;Zheng, Xin
    • Asian-Australasian Journal of Animal Sciences
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    • v.28 no.4
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    • pp.573-583
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    • 2015
  • B-32 is one of a panel of monoclonal anti-idiotypic antibodies to growth hormone (GH) that we developed. To characterize and identify its potential role as a novel growth hormone receptor (GHR) agonist, we determined that B-32 behaved as a typical $Ab2{\beta}$ based on a series of enzyme-linked immunosorbent assay assays. The results of fluorescence-activated cell sorting, indirect immunofluorescence and competitive receptor binding assays demonstrated that B-32 specifically binds to the GHR expressed on target cells. Next, we examined the resulting signal transduction pathways triggered by this antibody in primary porcine hepatocytes. We found that B-32 can activate the GHR and Janus kinase (2)/signal transducers and activators of transcription (JAK2/STAT5) signalling pathways. The phosphorylation kinetics of JAK2/STAT5 induced by either GH or B-32 were analysed in dose-response and time course experiments. In addition, B32 could also stimulate porcine hepatocytes to secrete insulin-like growth factors-1. Our work indicates that a monoclonal anti-idiotypic antibody to GH (B-32) can serve as a GHR agonist or GH mimic and has application potential in domestic animal (pig) production.

Effect of Samultanggamibang of Apoptosis of Melanoma cell (사물탕(四物湯) 가미방(加味方)이 흑색중(黑色腫) 세포고사(細胞枯死)에 미치는 효과(效果))

  • Park, Eun-Jung;Lee, Hai-Ja;Chang, Sung-Jin
    • The Journal of Pediatrics of Korean Medicine
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    • v.20 no.1
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    • pp.257-272
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    • 2006
  • Objective : In this study, the ability of Oriental medicine Samultanggamibang(SMTG) to induce apoptosis was investigated in B16F10 melanoma cells. Method : Tetrazolium-based colorimetric assay was performed for cytotoxicity test. Several new assays for the basis of biochemical events associated with apoptosis such as DNA fragmentation by a flow cytometry, caspase-3 activation and PARP cleavage by Western blotting should be carried out potentially useful for the basis of biochemical events associated with apoptosis such as a flow cytometry and caspase-3 activation. Results : (1) The number of B16F10 melanoma cells was less than 30 % after exposure to 1 mg/ml SMTG for 48 h. SMTG increased cytotoxicity of B16F10 melanoma cells in a dose- and time-dependent manner. (2) The percentage of apoptotic cells by flow cytometric analysis of the DNA-stained cells increased to 21 % at 24 h and 25 % at 48 h after treatment with 1 mg/ml SMTG. (3) SMTG-induced apoptosis was accompained by the activation of caspase-3 and the specific proteolytic cleavage of poly-ADP-ribose polymerase. (4) SMTG induces the activation of caspase-3 and the specific proteolytic cleavage of poly-ADP-ribose polymearse and eventually leads to apoptosis through c-Jun NH2-terminal protein kinase (JNK)-dependent manner in B16F10 melanoma cells. Conclusion : SMTG had a strong cytotoxic effect of B16F10 melanoma cells.

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Mechanism of Inhibition of HepG2 Cell Proliferation by a Glycoprotein from Hizikia fusiformis (톳(Hizikia fusiformis) 당단백질에 의한 HepG2 세포 증식 억제기전)

  • Ryu, Jina;Hwang, Hye-Jung;Kim, In-Hye;Nam, Taek-Jeong
    • Korean Journal of Fisheries and Aquatic Sciences
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    • v.45 no.6
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    • pp.553-560
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    • 2012
  • Hizikia fusiformis, a brown alga that is widely consumed in Korea, Japan, and China, possesses a number of potentially beneficial compounds, including antioxidants and anticoagulants. However, the molecular mechanisms of H. fusiformis in hepatoma cells have not been elucidated. This study investigated the antiproliferative effect and mechanism of action of a glycoprotein from H. fusiformis (HFGP) in HepG2 human hepatoma cells. In an MTS assay, 25 ${\mu}g/mL$ HFGP inhibited the proliferation of HepG2 cells by $52.36{\pm}2.37%$. HFGP caused the dose-dependent growth inhibition of HepG2 cells by inducing apoptosis and a sub-G1 phase arrest. The antiproliferative activity of HFGP was confirmed based on the expression of several apoptosis-related proteins, which was assessed by Western blot analysis. The expressions of Fas, Fas-associated death domain protein, Bax, and Bad was significantly up-regulated in HFGP-treated cells, and HFGP induced the translocation of Bax to mitochondria and the release of cytochrome c into the cytosol. Therefore, HFGP might be useful in the treatment of liver cancer.