• Food Science and Preservation
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• v.19 no.1
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• pp.123-130
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• 2012

In this study, 80% ethanolic extracts of tartary and common buckwheats were assessed for their total phenol content, total flavonoids content, antioxidant activity (DPPH, ABTS radical scavenging activity and reducing power), and anti-adipogenic effects. Our results show that total phenol contents of 80% ethanolic extract from tartary and common buckwheats were $17.35{\pm}0.41$ and $8.20{\pm}0.28\;{\mu}g$ GAE/g, respectively. Antioxidant activities of 80% ethanolic extract from tartary buckwheat were significantly higher than that of common buckwheat extract (p<0.05). During adipocyte differentiation, 80% ethanolic extracts of tartary and common buckwheat significantly inhibited lipid accumulation compared to control cells. We further evaluated the effect of buckwheat extracts on the changes of key gene expression associated with 3T3-L1 adipogenesis and ROS production. Tartary buckwheat extract was more suppressed the mRNA expressions ($PPAR{\gamma}$ and aP2) than that of common buckwheat extract. Moreover, tartary buckwheat inhibited the mRNA expression of both NOX4 (NADPH oxidase 4) and G6PDH (glucose-6-phosphate dehydrogenase). These results indicate that anti-adipogenesis effect of tartary buckwheat can be attributed to phenolic compound that may potentially inhibit ROS production.

• Tuberculosis and Respiratory Diseases
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• v.46 no.2
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• pp.185-194
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• 1999

Background: NF-$\kappa$B is a characteristic transcriptional factor whose functional activity is determined by post-translational modification of protein and subsequent change of subcellular localization. The involvement of the NF-$\kappa$B family of the transcription factors in the control of such vital cellular functions as immune response, acute phase reaction, replication of certain viruses and development and differentiation of cells has been clearly documented in many previous studies. Several recent observations have suggested that the NF-$\kappa$B might also be involved in the carcinogenesis of some hematological and solid tumors. Investigating the possibility that members of the NF-$\kappa$B family participate in the molecular control of malignant cell transformation could provide invaluable information on both molecular pathogenesis and cancer-related gene therapy. Method: To determine the expression patterns and functional roles of NF-$\kappa$B family transcription factors in human lung cancer cell lines NCI-H792, NCI-H709, NCI-H226 and NCI-H157 were analysed by western blot, using their respective antibodies. The nuclear and the cytoplasmic fraction of protein extract of these cell lines were subsequently obtained and NF-$\kappa$B expression in each fraction was again determined by western blot analysis. The type of NF-$\kappa$B complex present in the cells was determined by immunoprecipitation. To detect the binding ability of cell-line nuclear extracts to the KB consensus oligonucleotide, electrophoretic mobility shift assay(EMSA) was performed. Results: In the cultured human lung cancer cell lines tested, transcription factors of the NF-$\kappa$B family, namely the p50 and p65 subunit were expressed and localized in the nuclear fraction of the cellular extract by western blot analysis and immunocytochemistry. Immunoprecipitation assay showed that in the cell, the p50 and p65 subunits made NF-$\kappa$B complex. Finally it was shown by Electrophoretic Mobility Shift Assay(EMSA) that nuclear extracts of lung cancer cell lines are able to bind to NF-$\kappa$B consensus DNA sequences. Conclusion: These data suggest that in human lung cancer cell lines the NF-$\kappa$B p50/p65 complex might be activated. and strengthen the hypothesis that NF-$\kappa$B family transcription factors might be involved in the carcinogenesis of human lung cancer.

• Tuberculosis and Respiratory Diseases
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• v.55 no.1
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• pp.21-30
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• 2003

Background : Mucin synthesis in airways has been reported to be regulated by the epidermal growth factor receptor (EGFR) system. Epidermal growth factor receptor transactivation was identified as a critical element in G-protein-coupled receptors (GPCRs)-induced mitogenic signaling. EGF receptor transactivation by G-protein-coupled receptors requires metalloproteinase cleavage of proHB-EGF. This study was hypothesized that lipopolysaccharide (LPS)-induced mucin production associates with epidermal growth factor receptor transactivation, and MUC5AC production associates with epidermal growth factor receptor transactivation by G-protein-coupled receptors that regulates by metalloproteinase. Method : MUC5AC mucin production was examined in NCI-H292 cells and MUC5AC protein synthesis was assessed using ELISA. For the evaluation of mechanism of LPS-induced MUC5AC production, $TNF{\alpha}$ was measured using ELISA with or without pretreatment of heterotrimeric G-protein inhibitor, mastoparan. MUC5AC protein was measure with pretreatment of polyclonal $TNF{\alpha}$ antibody or mastoparan on LPS-induced MUC5AC production. For the evaluation of relation of G-protein and MUC5AC production, G-protein stimulant, mastopara-7, or matrix metalloproteinase, ADAM10, was added to NCI-H292 cells. MUC5AC protein was measure with pretreatment of polyclonal EGF antibody on mastoparan-7-induced MUC5AC production. Results : LPS alone did not increase significantly MUC5AC production. LPS with $TNF{\alpha}$ induced dose-dependently MUC5AC production in NCI-H292 cells. LPS increased dose-dependently $TNF{\alpha}$ secretion, which was inhibited by mastoparan. LPS with $TNF{\alpha}$-induced MUC5AC production was inhibited by neutralizing polyclonal $TNF{\alpha}$ antibody, mastoparan or AG 1472. Mastoparan-7 or ADAM10 increased dose-dependently MUC5AC production, which was inhibited by polyclonal neutralizing EGF antibody. Conclusion : In LPS-induced MUC5AC synthesis, LPS causes $TNF{\alpha}$ secretion, which induces EGFR expression. EGFR tyrosine kinase phosphorylation result in MUC5AC production. EGF-R transactivation by G-protein-coupled receptors requires matrix metalloproteinase cleavage of proHB-EGF.

• Molecules and Cells
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• v.37 no.2
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• pp.178-186
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• 2014

Differential transcription of the clusterin (CLU) gene yields two CLU isoforms, a nuclear form (nCLU) and a secretory form (sCLU), which play crucial roles in prostate tumorigenesis. Pro-apoptotic nCLU and anti-apoptotic sCLU have opposite effects and are differentially expressed in normal and cancer cells; however, their regulatory mechanisms at the transcriptional level are not yet known. Here, we examined the transcriptional regulation of nCLU in response to hypoxia. We identified three putative hypoxia response elements (HREs) in the human CLU promoter between positions -806 and +51 bp. Using a luciferase reporter, electrophoretic gel mobility shift, and chromatin immunoprecipitation assays, we further showed that hypoxia-inducible factor-$1{\alpha}$ (HIF-$1{\alpha}$) bound directly to these sites and activated transcription. Exposure to the hypoxia-mimetic compound $CoCl_2$, incubation under 1% $O_2$ conditions, or overexpression of HIF-$1{\alpha}$ enhanced nCLU expression and induced apoptosis in human prostate cancer PC3M cells. However, LNCaP prostate cancer cells were resistant to hypoxia-induced cell death. Methylation-specific PCR analysis revealed that the CLU promoter in PC3M cells was not methylated; in contrast, the CLU promoter in LNCap cells was methylated. Co-treatment of LNCaP cells with $CoCl_2$ and a demethylating agent promoted apoptotic cell death through the induction of nCLU. We conclude that nCLU expression is regulated by direct binding of HIF-$1{\alpha}$ to HRE sites and is epigenetically controlled by methylation of its promoter region.

• Ecology and Resilient Infrastructure
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• v.5 no.3
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• pp.163-173
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• 2018

The biopolymer (BP) used in this study is mainly composed of xanthan gum and ${\beta}$-glucan derived from microorganism and has been introduced as a novel material for soil stabilization. However, the broad applicability of BP has been suggested in the field of geotechnical engineering while little information is available about the effects of BP on the vegetation. The goal of this study is to find the BP effects on the growth of Camelina sativa L. (Camelina) under drought condition. For more thorough evaluation of BP effects on the plant growth, we examined not only morphological but also physiological traits and gene expression patterns. After 25 days of drought treatment from germination in the soil amended with 0, 0.25, 0.5, and 1% BP, we observed that the BP concentration was strongly correlated the growth of Camelina. When plants were grown under drought stress, Camelina in 0.5% BP mixture showed better physiological parameters of the leaf stomatal conductance, electrolyte leakage and relative water content compared to those in control soil without BP. Plant recovery rate after re-watering was higher and the development of lateral root was lower in BP amended soil. RNA expression of Camelina leaf treated with/without drought for 7 and 10 days showed that aquaporin genes transporting solutes at bio-membrane, CsPIP1;4, 2;1, 2;6 and TIP1;2, 2;1, were induced more in the plants with BP amendment and drought treatment. These results suggest that the soil amended with BP has a positive effect on the transport of nutrients and waters into Camelina by improving water retention in soil under drought condition.

• Applied Microscopy
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• v.34 no.3
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• pp.171-178
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• 2004

Metallothionein (MT) is a family of ubiquitous, low molecular weight (6-7 kDa), cysteine-rich protein with a high affinity to metal ions and has no aromatic amino acids and histidine. Some of the known functions of MT include detoxification of heavy metals and alkylating agents and neutralization of free radicals. Also, this protein may affect a number of cellular processes including gene expression, apoptosis, proliferation and differentiation. But, its actual functions are still not clear. The present study was undertaken to examine immunocytochemically the localization of MT in developing rat liver. On the day 11 of gestation, the fetal rat liver has already been formed and contained numerous oval cells with high nuclear cytoplasmic ratio, which were the progenitors of hepatic parenchymal cells, but no reaction products of MT were detected at this time. And then, positive reactions against MT started to appear predominantly in the parenchymal cells of liver from the 13th day after gestation. Reaction products, immunogold particles or brown coloration, were localized at both the nucleus and the cytoplasm of the parenchymal cells, except mitochondria. The intensity of this reaction gradually increased, and exhibited the strongest at birth. The intensity of MT staining and immunogold labelling diminished with growth, and by the 15th day after birth weak positive reaction was observed in the cells. In brief, positive reactions for MT were observed in the oval cells and the parenchymal cells during fetal stage, meanwhile they were present only in the parenchymal cells after birth. The present results suggest that MT possibly involves parechymal cell proliferation and differentiation through the storage or the supply of various metal ions in the developing rat liver.

• Radiation Oncology Journal
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• v.19 no.4
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• pp.389-396
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• 2001

Purpose : To detect differentially expressed genes in the patients with uterine cervical cancer during the radiation therapy. Materials and Methods : In patients with biopsy proven uterine cervical cancer, we took tumor tissue just before radiation therapy and at 40 minutes after external irradiation of 1.8 Gy. Total RNAs isolated from non-irradiated and irradiated tumor tissue samples were analyzed using the differential-display reverse transcription-polymerase chain reaction (DDRT-PCR). Complementary DNA (cDNA) fragments corresponding to differentially expressed messenger RNAs(mRNAs) were eluted, and cloned. The differential expression of the corresponding mRNAs was confirmed by reverse northern blot. Differentially expressed cDNA bands were sequenced. Nucleotide sequence data were analyzed in the Gene Bank and EMBL databases via the BLAST network sewer to identify homologies to known genes or cDNA fragments. Expression pattern of down-regulated clone was examined using RT-PCR in S patients undergoing radiotherapy. Results : We identified 18 differentially expressed bands by DDRT-PCR, which were eluted and cloned. There were 10 up-regulated clones and 1 down-regulated clone in reverse northern blot. One cDNA fragment had homology to chemokine receptor CXCR4, four were identified as Human ESTs in the EMBL database in EST clones. Down-regulated CxCa-11 was also down regulated in all patients. Conclusion : Using the DDRT-PCR, we have identified 10 up-regulated and 1 down-regulated clone(s) in the patients with uterine cervical cancer during the radiation therapy. The clinical relevance and the functions of these genes will be further investigated.

• Journal of Periodontal and Implant Science
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• v.37 no.sup2
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• pp.447-463
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• 2007

Periodontal ligament (PDL) cells have been known as multipotential cells, and as playing an important rolesin periodontal regeneration. The PDL cells are composed of heterogeneous cell populations which have the capacity to differentiate into either cementoblasts or osteoblasts, depending on needs and conditions. Therefore, PDL cells have the capacity to produce mineralized nodules in vitro in mineralization medium which include ascorbic acid, ${\beta}$-glycerophosphate and dexamethasone. In spite of these well-known osteoblast like properties of PDL cells, very little is known about the molecules involved in the formation of the mineralized nodules in the PDL cells. In the present study, we analysed gene-expression profiles during the mineralization process of cultured PDL cells by means of a cDNA microarray consisting of 3063 genes. Nodules of mineralized matrix were strongly stained with alizarin red S on the PDL cells cultured in the media with mineralization supplements. Among 3,063 genes analyzed, 35 were up-regulated more than two-fold at one or more time points in cells that developed matrix mineralization nodules, and 38 were down-regulated to less than half their normal level of expression. In accord with the morphological change we observed, several genes related to calcium-related or mineral metabolism were induced in PDL cells during osteogenesis, such as IGF-II and IGFBP-2. Proteogycan 1, fibulin-5, keratin 5, ,${\beta}$-actin, ${\alpha}$-smooth muscle actin and capping protein, and cytoskeleton and extracellular matrix proteins were up-regulated during mineralization. Several genes encoding proteins related to apoptosis weredifferentially expressed in PDL cells cultured in the medium containing mineralization supplements. Dkk-I and Nip3, which are apoptosis-inducing agents, were up-regulated, and Btf and TAXlBP1, which have an anti-apoptosis activity, were down-regulated during mineralization. Also periostin and S100 calciumbinding protein A4 were down-regulated during mineralization.

• KOREAN JOURNAL OF CROP SCIENCE
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• v.56 no.3
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• pp.264-272
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• 2011

It needs to develop high phosphate-available rice that is able to minimize environmental pollution caused by phosphate fertilizer. Then we have transformed 4 rice transporter genes, OsPT(Oryza sativa Phosphate Transporter)1, OsPT4, OsPT7 or OsPT8, to rice (Oryza sativa cv. Dongjin) via Agrobacterium-mediated transformation. We tested adaptation in the P-deficient condition of Dongjin (parental) and each transgenic line in the pot and the field conditions. Definite physiological changes have been observed in OsPTs transgenic lines including culm length, root formation and heading date. Phosphate uptake at harvesting stage was about three times higher in OsPT1-OX (overexpression) and OsPT4-OX than in Dongjin (wt) without P application. There are no variations in total phosphate-content of brown rice of OsPT1-OX in spite of high phosphate uptake. Practically the expression of OsPT1 has contributed to stabilize grain production without P fertilization in rice cultivation than Dongjin.

• Journal of Life Science
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• v.17 no.2 s.82
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• pp.198-203
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• 2007

DNA transfection is a powerful tool for studying gene functions. The $Ca^{2+}$-phosphate precipitation remains one of the most popular and cost-effective transfection techniques. Mature neurons are more resistant to transfection than young ones and most other cell types, and easy to die if microenvironment changes. Here, we report a transfection protocol for mature neurons. The critical modifications are inclusion of glial cells in culture and careful control of $Ca^{2+}$-phosphate precipitation under microscope. Cerebral glial cells were grown until ${\sim}70-80%$ confluence in DMEM/10% horse serum, which was thereafter replaced with serum-free Neurobasal/Ara-C, and 319 hippocampal neurons were plated onto the glial layer Formation of fine $DNA/Ca^{2+}$-phosphate precipitates was induced using Clontech $CalPhos^{TM}$ Mammalian Transfection Kit, and the size ($0.5-1\;{\mu}m$ in diameter) and density(about 10 particles/$100\;{\mu}m^2$) were carefully controlled by the time of incubation in the medium. This modified protocol can be reliably applied for transfection of mature neurons that are maintained longer than two weeks in vitro, resulting in 10-15 healthy transfected neurons per a well of 24-well plates. The efficacy of the protocol was verified by punctate expression of $pEGFP-CaMKII{\alpha}$, a synaptic protein, and diffuse expression of pDsRed2. Our protocol provides a reliable method for transfection of mature neurons in vitro.