• Fisheries and Aquatic Sciences
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• v.21 no.3
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• pp.8.1-8.6
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• 2018

Desmarestia japonica (Phaeophyceae, Desmarestiales) was recently established from the Japanese ligulate Desmarestia and is morphologically similar to D. ligulata. This species has been reported only from Japan. However, the taxonomic reports based on additional regional distributions are needed to clarify this taxonomic entity and its species boundaries. Because Desmarestia species have restricted distributions in Korea, we reexamined herbarium specimens of D. ligulata deposited at the National Institute of Biological Resources (South Korea). To improve the amplification efficiency of the polymerase chain reaction and avoid contamination by the DNA of other organisms, we developed taxon-specific molecular markers suitable for DNA barcoding of Desmarestia species. Nuclear ribosomal small subunit RNA (18S rDNA) and mitochondrial cytochrome c oxidase 1 (cox1) regions were selected as target DNA. As a result, both were successfully isolated from herbarium specimens of D. japonica acquired over 10 years. These molecular markers provide useful genetic information for herbarium specimens for which conventional molecular analysis is challenging.

• Molecules and Cells
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• v.41 no.4
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• pp.290-300
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• 2018

Using an in vitro model of intestinal organoids derived from intestinal crypts, we examined effects of indole-3-carbinol (I3C), a phytochemical that has anticancer and aryl hydrocarbon receptor (AhR)-activating abilities and thus is sold as a dietary supplement, on the development of intestinal organoids and investigated the underlying mechanisms. I3C inhibited the in vitro development of mouse intestinal organoids. Addition of ${\alpha}$-naphthoflavone, an AhR antagonist or AhR siRNA transfection, suppressed I3C function, suggesting that I3C-mediated interference with organoid development is AhR-dependent. I3C increased the expression of Muc2 and lysozyme, lineage-specific genes for goblet cells and Paneth cells, respectively, but inhibits the expression of IAP, a marker gene for enterocytes. In the intestines of mice treated with I3C, the number of goblet cells was reduced, but the number of Paneth cells and the depth and length of crypts and villi were not changed. I3C increased the level of active nonphosphorylated ${\beta}$-catenin, but suppressed the Notch signal. As a result, expression of Hes1, a Notch target gene and a transcriptional repressor that plays a key role in enterocyte differentiation, was reduced, whereas expression of Math1, involved in the differentiation of secretory lineages, was increased. These results provide direct evidence for the role of AhR in the regulation of the development of intestinal stem cells and indicate that such regulation is likely mediated by regulation of Wnt and Notch signals.

• Molecules and Cells
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• v.45 no.6
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• pp.403-412
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• 2022

Hypoxia leads to significant cellular stress that has diverse pathological consequences such as cardiovascular diseases and cancers. MicroRNAs (miRNAs) are one of regulators of the adaptive pathway in hypoxia. We identified a hypoxia-induced miRNA, miR-34c, that was significantly upregulated in hypoxic human umbilical cord vein endothelial cells (HUVECs) and in murine blood vessels on day 3 of hindlimb ischemia (HLI). miR-34c directly inhibited BCL2 expression, acting as a toggle switch between apoptosis and autophagy in vitro and in vivo. BCL2 repression by miR-34c activated autophagy, which was evaluated by the expression of LC3-II. Overexpression of miR-34c inhibited apoptosis in HUVEC as well as in a murine model of HLI, and increased cell viability in HUVEC. Importantly, the number of viable cells in the blood vessels following HLI was increased by miR-34c overexpression. Collectively, our findings show that miR-34c plays a protective role in hypoxia, suggesting a novel therapeutic target for hypoxic and ischemic diseases in the blood vessels.

• Asian Pacific Journal of Cancer Prevention
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• v.16 no.9
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• pp.3961-3968
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• 2015

Background: Variants of human papillomavirus (HPV) show more oncogenicity than do prototypes. The HPV16 Asian variant (HPV16As) plays a major role in cervical cancer of Asian populations. Some amino acid changes in the E6 protein of HPV16 variants affect E6 functions such as p53 interaction and host immune surveillance. This study aimed to investigate activities of HPV16As E6 protein on modulation of expression of miRNA-21 as well as interferon regulatory factors (IRFs) 1, 3, 7 and c-fos. Materials and Methods: Vectors expressing E6 protein of HPV16As (E6D25E) or HPV16 prototype (E6Pro) were constructed and transfected into C33A cells. HCK1T cells expressing E6D25E or E6Pro were established by transducing retrovirus-containing E6D25E or 16E6Pro. The E6AP-binding activity of E6 and proliferation of the transfected C33A cells were determined. MiR-21 and mRNA of interesting genes were detected in the transfected C33A cells and/or the HCK1T cells, with or without treatment by culture medium from HeLa cells (HeLa-CM). Results: E6D25E showed binding activity with E6AP similar to that of E6Pro. Interestingly, E6D25E showed a higher activity of miR-21 induction than did E6Pro in C33A cells expressing E6 protein. This result was similar to the HCK1T cells expressing E6 protein, with HeLa-CM treatment. The miR-21 up-regulation significantly corresponded to its target expression. Different levels of expression of IRFs were also observed in the HCK1T cells expressing E6 protein. Interestingly, when treated with HeLa-CM, IRFs 1, 3 and 7 as well as c-fos were significantly suppressed in the HCK1T cells expressing E6D25E, whereas those in the HCK1T cells expressing E6Pro were induced. A similar situation was seen for IFN-${\alpha}$ and IFN-${\beta}$. Conclusions: E6D25E of the HPV16As variant differed from the E6 prototype in its activities on epigenetic modulation and immune surveillance and this might be a key factor for the important role of this variant in cervical cancer progression.

• Entomological Research
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• v.48 no.5
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• pp.384-389
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• 2018

Aphis gossypii Glover is an important insect pest that functions as a viral vector and mediates approximately 45 different viral diseases. As part of a strategy for control of A. gossypii, we investigated the functions of genes using RNAi. To this end, a cDNA library was constructed for various genes and for selecting appropriate targets for RNAi mediated silencing. The cDNA library was constructed using the Gateway cloning system with site-specific recombination of bacteriophage ${\lambda}$. It was used to carry out single step cloning of A. gossypii cDNAs. As a result, a cDNA library with a titer of $8.4{\times}10^6$ was constructed. Since the sequences in this library carry att sites, they can be cloned into various binary vectors. This library will be of value for various studies. For later screening of selected genes, it is planned to clone the library into virus-induced gene silencing (VIGS) vectors, which makes it possible to analyze gene function and allow subsequent transfection of plants. Such transfection experiments will allow testing of RNAi-induced insecticidal activity or repellent activity to A. gossypii, and result in the identification of target genes. It is also expected that the constructed cDNA library will be useful for analysis of gene functions in A. gossypii.

• Korean Journal of Head & Neck Oncology
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• v.21 no.2
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• pp.158-164
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• 2005

Purpose: The serine/threonine kinase Akt was described to inhibit apoptosis in cancer. This study was to examine the effect of Gleevec on head and neck squamous cell carcinoma(HNSCC) through the mechanism of Akt. Experimental Design: Gleevec was introduced into the HNSCC cell lines UMSCC10B, HN12 and HN30 in a range of concentrations. Cell viability was assessed by clonogenic survival analysis. Targets of Gleevec(PDGFR, c-Kit, and c-Abl) were evaluated by Western blot. HNSCC tissue samples were stained for PDGFR, c-Kit and phosphorylated Akt. Akt phosphorylation following Gleevec treatment was assessed using Western blot. Akt siRNA was used to as the positive control. Results: Colony forming efficiency decreased with an increase in concentration of Gleevec. Expressions of PDGFR, c-Kit, and c-Abl were observed in HNSCC cells. Immunohistochemistry confirmed high expression of PDGFR, c-Kit, and p-Akt in human HNSCC tissues. Akt kinase activity was significantly inhibited with increasing concentration of Gleevec in HNSCC cells, and near complete dephosphorylation of Akt was observed at $6{\mu}M$ of Gleevec in the UMSCC10B and HN30 cell lines. Conclusions: Gleevec at clinically comparable concentrations caused a dose dependant decrease in HNSCC survival. The decreased cell survival was related to the inhibition of Akt kinase activity and dephosphorylation of Akt. Akt signaling pathway may be a relevant target for Gleevec in treating HNSCC.

• Korean Journal of Microbiology
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• v.54 no.2
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• pp.149-151
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• 2018

Clostridium perfringens is a Gram-positive, rod-shaped, anaerobic, spore-forming pathogenic bacterium, which belongs to the Clostridiaceae family. C. perfringens causes diseases including food poisoning in vertebrates and intestinal tract of humans. Bacteriophages that can kill target bacteria specifically have been considered as one of control methods for bacterial pathogens. Here, we report a draft genome sequence of the bacteriophage CP3 effective to C. perfringens. The phage genome comprises 52,068 bp with a G + C content of 34.0%. The draft genome has 74 protein-coding genes, 29 of which have predicted functions from BLASTp analysis. Others are conserved proteins with unknown functions. No RNAs were found in the genome.

• Asian-Australasian Journal of Animal Sciences
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• v.31 no.8
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• pp.1366-1372
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• 2018

Objective: A disintegrin and metallopeptidase with thrombospondin motifs type 8 (ADAMTS8) is crucial for diverse physiological processes, such as inflammation, tissue morphogenesis, and tumorigenesis. The chicken ADAMTS8 (chADAMTS8) gene was differentially expressed in the kidney following exposure to different calcium concentrations, suggesting a pathological role of this protein in metabolic diseases. We aimed to examine the molecular characteristics of chADAMTS8 and analyze the gene-expression differences in response to toll-like receptor 3 (TLR3) stimulation. Methods: The ADAMTS8 mRNA and amino acid sequences of various species (chicken, duck, cow, mouse, rat, human, chimpanzee, pig, and horse) were retrieved from the Ensembl database and subjected to bioinformatics analyses. Reverse-transcription polymerase chain reaction (RT-PCR) and quantitative PCR (qPCR) experiments were performed with various chicken tissues and the chicken fibroblast DF-1 cell line, which was stimulated with polyinosinic-polycytidylic acid (poly[I:C]; a TLR3 ligand). Results: The chADAMTS8 gene was predicted to contain three thrombospondin type 1 (TSP1) domains, whose amino acid sequences shared homology among the different species, whereas sequences outside the TSP1 domains (especially the amino-terminal region) were very dif­ferent. Phylogenetic analysis revealed that chADAMTS8 is evolutionarily clustered in the same clade with that of the duck. chADAMTS8 mRNA was broadly expressed in chicken tissues, and the expression was significantly up-regulated in the DF-1 cells in response to poly(I:C) stimulation (p<0.05). These results showed that chADAMTS8 may be a target gene for TLR3 signaling. Conclusion: In this report, the genetic information of chADAMTS8 gene, its expression in chicken tissues, and chicken DF-1 cells under the stimulation of TLR3 were shown. The result suggests that chADAMTS8 expression may be induced by viral infection and correlated with TLR3-mediated signaling pathway. Further study of the function of chADAMTS8 during TLR3-dependent inflammation (which represents RNA viral infection) is needed and it will also be important to examine the molecular mechanisms during different regulation, depending on innate immune receptor activation.

• Molecules and Cells
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• v.41 no.5
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• pp.423-435
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• 2018

This investigation was aimed at working out the combined role of lncRNA H19, miR-29b and Wnt signaling in the development of colorectal cancer (CRC). In the aggregate, 185 CRC tissues and corresponding para-carcinoma tissues were gathered. The human CRC cell lines (i.e. HT29, HCT116, SW480 and SW620) and normal colorectal mucosa cell line (NCM460) were also purchased. Si-H19, si-NC, miR-29b-3p mimics, miR-29b-3p inhibitor, si-PGRN and negative control (NC) were, respectively, transfected into the CRC cells. Luciferase reporter plasmids were prepared to evaluate the transduction activity of $Wnt/{\beta}-catenin$ signaling pathway, and dual-luciferase reporter gene assay was arranged to confirm the targeted relationship between H19 and miR-29b-3p, as well as between miR-29b-3p and PGRN. Finally, the proliferative and invasive capacities of CRC cells were appraised through transwell, MTT and scratch assays. As a result, overexpressed H19 and down-expressed miR-29b-3p displayed close associations with the CRC patients' poor prognosis (P < 0.05). Besides, transfection with si-H19, miR-29b-3p mimic or si-PGRN were correlated with elevated E-cadherin expression, decreased snail and vimentin expressions, as well as less-motivated cell proliferation and cell metastasis (P < 0.05). Moreover, H19 was verified to directly target miR-29b-3p based on the luciferase reporter gene assay (P < 0.05), and miR-29b-3p also bound to PGRN in a direct manner (P < 0.05). Finally, addition of LiCl ($Wnt/{\beta}-catenin$ pathway activator) or XAV93920 ($Wnt/{\beta}-catenin$ pathway inhibitor) would cause remarkably altered E-cadherin, c-Myc, vimentin and snail expressions, as well as significantly changed transcriptional activity of ${\beta}-catenin/Tcf$ reporter plasmid (P < 0.05). In conclusion, the lncRNA H19/miR-29b-3p/PGRN/Wnt axis counted a great deal for seeking appropriate diagnostic biomarkers and treatment targets for CRC.

• YAKHAK HOEJI
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• v.59 no.5
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• pp.207-214
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• 2015

P2Y receptors, a type of P2 receptor family, are G-protein coupled receptors and 8 subtypes have been characterized ($P2Y_1$, $P2Y_2$, $P2Y_4$, $P2Y_6$, $P2Y_{11-14}$). Recently, several studies have shed light on the role of P2Y receptors in bone biology. Among them, little is known on the role of $P2Y_6$ receptor on osteoclast differentiation. Thus, we investigated the role of $P2Y_6$ receptor on osteoclastogenesis using $P2Y_6$ receptor selective antagonist, MRS 2578. When osteoblasts and bone marrow cells were co-cultured in the presence of $VitD_3$ and $PGE_2$, $P2Y_6$ antagonist increased the formation of TRAP positive osteoclasts. To elucidate the target cells of $P2Y_6$ antagonist, we first checked the effect of MRS 2578 on osteoblasts. Treatment of MRS 2578 did not affect OPG : RANKL mRNA ratio in osteoblasts. Next, we checked the effects of $P2Y_6$ antagonist on osteoclast precursors using mouse bone marrow macrophages (BMMs). Addition of MRS 2578 increased the number of osteoclasts in RANKL-treated BMMs. Although $P2Y_6$ antagonist had no effect on RANKL-induced NFATc1, c-Fos and MafB expression levels, it significantly stimulated RANKL-induced Blimp1 mRNA expression in BMMs. Taken together, these data indicate that $P2Y_6$ antagonist increases osteoclast formation by upregulation of Blimp1 expression.