• Journal of Acupuncture Research
• /
• v.19 no.2
• /
• pp.28-38
• /
• 2002

Objective : This study is aimed to investigate the effects of bee venom and purified bee venom on cell death in synovial cell line. Methods : It was evaluated by using MTT assay, morphological method, flow cytometry, immunocytochemistry analysis, RT-PCR. Results : The result obtained is as follows. 1. The MTT assay demonstrated that synovial cell viability was significantly inhibitted dose-dependently by treatment with BV and PBV in comparison with control. And the inhibitory effect of BV and PBV was almost same. 2. The morphologic study demonstrated that synovial cell showed apoptotic body resulted from apoptosis after treatment with BV and PBV for 6 hours using microscope. 3. The Flow cytometry demonstrated that apoptosis of synovial cell treated with BV and PBV was related with stop of cell cycle in stage of G0/G1. 4. Immunocytochemistry assay demonstrated that COX-II and iNOS were slightly expressed by treatment with BV and PBV in comparison with control group. 5. RT-PCR analysis demonstrated that COX-II were almost down-regulated by high dose treatment with BV and PBV in comparison with control group. iNOS were well down-regulated by treatment with $5{\mu}g/ml$ BV and PBV whereas it was well expressed in control group. Conclusion : These results suggest that bee venom and purified bee venom have significant effect on cell death in synovial cell line and further study is needed in vivo.

• The Korea Journal of Herbology
• /
• v.31 no.5
• /
• pp.47-53
• /
• 2016

Objectives : Mori Follium (Morus alba L. leaf) has been cultivated in many Asian countries. Especially, mulberry leaf has been used as an anti-diabetic remedy in oriental medicine. However, anti-obesity effect of mulberry has not been unknown. In this study, our objectives of study is to investigate the anti-adipogenic effect of mulberry water extract (MLE) and to reveal potential molecular anti-obesity mechanism in 3T3-L1 adipocytes differentiation model.Methods : The cytotoxicity of MLE in 3T3-L1 was examined by MTT assay. Anti-adipogenic effect of MLE was evaluated by Oil Red O (ORO) staining. To elucidate the molecular mechanism, inhibitor assay was employed. The mRNA expression levels of adipogenic transcriptional factors such as PPARγ and fatty acid synthase (FAS) were analyzed by reverse transcription-polymer chain reaction (RT-PCR) analysis.Results : The MLE treatment for 24 h did not affect to the 3T3-L1 cells at concentrations of 1, 10, 100, 200, 400, 800 and 1,000 ㎍/㎖. Thus, non-toxic concentration rages of MLE were used during adipogenesis period (day -2 to 7). Intracellular lipid accumulation in MLE-treated 3T3-L1 adipocytes (day 6) were quantitatively evaluated by ORO staining. The MLE treatment significantly and dose-dependently suppressed 3T3-L1 adipogenesis by 60.42%, 38.24%, and 5.97% at 10, 100, and 200 ㎍/㎖, respectively. In addition, our inhibitor assay and RT-PCR analysis revealed that the MLE-inhibited 3T3-L1 adipogenesis through inhibition of PPARγ mediated by Wnt/β-catenin signaling pathway.Conclusions : In conclusion, these findings indicate that the MLE could be used in prevent and/or treatment of obesity-related diseases.

• YAKHAK HOEJI
• /
• v.52 no.5
• /
• pp.376-383
• /
• 2008

Chungkookjang (CKJ) is a fermented soybean product and one of favorite traditional foods in Korea. In this study, the alcoholic extract from Korean fermented soybean (CKJ) and its one of major flavonoids, genistein were evaluated for their protective effect against B(a)P induced cytotoxicity and DNA damage in HepG2 cells. CKJ extract and genistein decreased B(a)P-induced cell cytotoxicity. CKJ extract inhibited DNA single strand breaks evaluated by single cell gel electrophoresis. From RT-PCR study, it was revealed that CKJ extract decrease DNA damage induced in HepG2 cells expressing CYP1A1 and 1A2 by B(a)P. The metabolizing activities of CYP1A1 and CYP1A2, as measured by the 7-alkoxy resorufin O-deethylation (AROD) assay, showed that CKJ extract and genistein inhibited CYP1A1 and CYP1A2 activities. Genistein may contribute to these biological effects of CKJ extract at least in part. All these results indicate that CKJ extract and genistein may be useful for protection against B(a)P-induced cytotoxicity and DNA damage. Therefore, the alcoholic extract of Korean fermented soybean (CKJ) is suggested to be promising functional food which can prevent the cellular genotoxicity of dietary and lifestyle related carcinogens.

• Journal of Acupuncture Research
• /
• v.19 no.1
• /
• pp.189-202
• /
• 2002

Objective : This study is designed to investigate the effects of bee venom and melittin on cell death in neuroblastoma cell line after pretreatment with NG(nerve growth inhibitory substance) Methods : It was evaluated by using MTT assay, morphological method, DNA fragmenation, flow cytometry, immunocytochemistry analysis, RT-PCR and Western blot. Results : The MTT assay demonstrated that neuroblastoma cell viability was significantly inhibited dose-dependently by treatment with bee venom and melittin after pretreatment with NG in comparison awith control. The morphological study and fow cytometry demonstrated that neuroblastoma cell showed apoptosis. DNA fragmenation showed DNA ladder below 1 Kbp. Immunocytochemistry assay demonstrated that Fos and MAPK were down-regulated. RT-PCR analysis demonstrated that Fos and MAPK was down-regulated. Western blot demonstrated that Fos and MAPK were down-regulated from $1{\mu}g/ml$ bee venom in neuroblastoma cell pretreated with NG. Conclusion : These result suggests that bee venom and melittin after NG treatment have significant anti-cancer effect and further study is needed in vivo.

• Ocean and Polar Research
• /
• v.41 no.2
• /
• pp.79-88
• /
• 2019

In this study, the inhibitory effect of Atriplex gmelinii C. A. Mey. against the activity of MMP-2 and MMP-9 secreted from phorbol-12-myristate-13-acetate (PMA)-stimulated HT-1080 cells was evaluated by gelatin zymography and enzyme-linked immunosorbent assay (ELISA), reverse transcription polymerase-chain reaction (RT-PCR), and Western blot assay. Specimens of the halophyte A. gmelinii were extracted twice for 24 hr with methylene chloride ($CH_2Cl_2$), and then twice with methanol (MeOH), in turn. Each extract significantly inhibited the enzymatic activities in gelatin zymography and MMP ELISA kit, and expression of MMP-2 and 9 in mRNA and protein levels. Two crude extracts were combined and then the combined crude extracts were fractionated into n-hexane, 85% aqueous methanol (85% aq.MeOH), n-butanol (n-BuOH), and water ($H_2O$) fractions, according to solvent polarity. Among solvent-partitioned fractions, the 85% aq.MeOH fraction showed the strongest inhibitory effect against MMP-2 and -9 in gelatin zymography and MMP ELISA kit. In RT-PCR, all solvent-partitioned fractions significantly suppressed mRNA expression of MMP-2 and -9. On the other hand, in Western blot assay, all solvent-partitioned fractions except $H_2O$ significantly reduced expression levels of protein. HT 1080 cell migration was most significantly inhibited by the n-BuOH fraction followed by the 85% aq.MeOH and $H_2O$ fractions. These results suggest that A. gmelinii could be used as a potential source to inhibit tumor cell metastasis.

• Ocean and Polar Research
• /
• v.40 no.4
• /
• pp.249-257
• /
• 2018

Matrix metalloproteinases (MMPs) are associated with the invasion and metastasis of malignant tumors composed of cancer cells in an increased state of expression. This study evaluates the inhibitory effect of Carex pumila on MMP-2 and MMP-9 activity in phorbol-12-myristate-13-acetate (PMA)-stimulated HT-1080 human fibrosarcoma cells using gelatin zymography, MMPs enzyme-linked immunosorbent assay (ELISA), reverse transcription-polymerase chain reaction (RT-PCR) and Western blot assay. C. pumila was extracted twice with dichloromethane ($CH_2Cl_2$) and methanol (MeOH). Treatment with $CH_2Cl_2$ extract and MeOH extract in PMA-stimulated HT-1080 cells effectively reduced the production of MMP-2 and 9. Also, the combined crude extracts ($CH_2Cl_2$ and MeOH) significantly inhibited the enzymatic activities and the expression of MMP-2 and MMP-9 in mRNA and protein levels. The combined crude extracts were partitioned between $CH_2Cl_2$ and water. The organic layer was further fractionated with n-hexane, 85% aqueous methanol (85% aq.MeOH) and the aqueous layer was separated into n-butanol and water, successively. Of the fractions, 85% aq.MeOH fraction showed the highest inhibitory activity of MMP-2 and MMP-9 in gelatin zymography and MMP ELISA kit. Furthermore, 85% aq.MeOH fraction most significantly suppressed cell migration. In RT-PCR and Western blot assay, n-butanol and 85% aq.MeOH fractions exerted the greatest inhibition on mRNA and protein expression of MMP-2 and MMP-9, respectively. As a result, C. pumila can be used as a good anti-invasive agent source.

• Journal of Plant Biotechnology
• /
• v.34 no.4
• /
• pp.375-380
• /
• 2007

This study was conducted to develop herbicide-resistance domestic maize plants by introducing the CP4 5-enol-pyruvylshikimate-3-phosphate synthase (CP4 EPSPS) gene using Agrobacterium tumefaciens-mediated immature embryo transformation. Immature embryos of five genotypes (HW1, KL103, HW3, HW4, HW7) were co-cultivated with strains Agrobacterium tumefaciens (strain C58C1) containing the binary vector (pCAMBIA2300) carrying Ubiquitin promoter-CP4 EPSPS gene and Cauliflower mosaic virus 35S (CaMV35S) promoter-nptll gene conferring resistance to paromomycin as a selective agent. The presence and expression of CP4 EPSPS transgene were confirmed by PCR, RT-PCR and Northern blot analysis, respectively. Also, the resistance to glyphosate in the transgenic maize ($T_1$) was analyzed by shikimate accumulation assay. The frequency (%) of paromomycin-resistance callus was 0.37, 0.03, 2.20, 2.37, and 0.81% in pure lines HW1, KL103, HW3, HW4 and HW7, respectively. EPSP transgene sequences were amplified in putative transgenic plants that regenerated from paromomycin-resistance calli of two inbred lines (HW3, HW4). Of them, RT-PCR and Northern blot analyses revealed that the transgene was only expressed in two transgenic events (M266, M104) of HW4 inbred line, and a mild glyphosate resistance of transgenic event (M266) was confirmed by the lower shikimate accumulation in leaf segments. These results demonstrate that transgenic maize with herbicide-resistance traits in Korean genotype can be genetically obtained.

• The Journal of Internal Korean Medicine
• /
• v.27 no.1
• /
• pp.166-177
• /
• 2006

Objectives : This study was performed to determine if Orostschys japonicus A. Berger has protective effects against CML in K562 cell lines. Materials and Methods : MTT assay, cell proliferation assay, Reverse transcription-polymerase reaction chain, RT-PCR, DNA fragmentation assay, Quantitative PCR were studied. Results : Orostschys japonicus A. Berger had no effects on Bax gene in K562 cell lines, but decreased Bcl-2 gene, and increased the Caspases-3 gene. This is indicate of induced apoptosis in K562 cell lines by Orostschys japonicus A. Berger. Conclusion : These results suggest that Orostschys japonicus A. Berger has effects on apatosis in K562 cell lines.

• Microbiology and Biotechnology Letters
• /
• v.45 no.2
• /
• pp.87-92
• /
• 2017

The first Ebola hemorrhagic fever outbreak occurred in the Democratic Republic of Congo and Sudan in 1976 and then emerged in West Africa in 2014 with a total of 27,741 cases and 11,284 deaths. The fever is caused by the Ebola virus, which belongs to the Filoviridae family and contains a ssRNA genome. The known subtypes of the virus are Bundibugyo ebolavirus, Reston ebolavirus, Sudan ebolavirus, $Ta\ddot{i}$ Forest ebolavirus, and Zaire ebolavirus. The Ebola outbreak was historically originated majorly from the East and Central African tropical belt. The current outbreaks in West Africa caused numerous deaths and spread fear in global society. In the absence of effective treatment strategies and any vaccine, accurate diagnosis is the most important contributing factor in the management and control of the epidemic disease. WHO (World Health Organization) has announced emergency guidance for the selection and use of Ebola in in vitro diagnostic assays. Numerous companies and research institutions have studied the various diagnosis methods and identified four WHO procurement approved as diagnosis kits: RealStar Ebolavirus Screen RT-PCR kit 1.0 (Altona), Liferiver-Ebola Virus (EBOV) Real time RT-PCR kit, Xpert Ebola Assay, and ReEBOV Antigen Rapid Test Kit. The efficiency of novel diagnostic kits such as Rapid Diagnosis Test (RDT) is currently being evaluated.

• Korean Journal of Veterinary Service
• /
• v.42 no.1
• /
• pp.31-37
• /
• 2019

There have been a total tenth FMD outbreaks in Korea and for the first time, type O and A were detected simultaneously in 2017, which led to difficulties in FMD control. For the effective prevention of FMD, the importance of discrimination of serotypes became greater. Therefore, the most urgent requirement in case of FMD outbreak is differential diagnosis of serotypes. In this study, we developed a PNA probe-mediated multiplex real-time reverse transcription-polymerase chain reaction (rRT-PCR) assay using the peptide nucleic acid (PNA) probe, which is known to be stable to nucleotide mutation and that could specifically detect the all FMDV serotype A, FMDVA Yeoncheon strain which was occurred in Korea in 2017, and FMDV A viruses shown 96% similarity with FMDVA/Yeoncheon strain, at the same time. Therefore, It is believed that the newly introduced FMDVA will be effectively diagnosed using the PNA probe multiplex RT-PCR developed in this study, and ultimately contribute to the prevention of FMD.