• Korean journal of applied entomology
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• 제56권1호
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• pp.19-28
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• 2017

Polydnaviruses (PDVs) are a group of double-stranded DNA viruses symbiotic to some endoparasitoid wasps. Cotesia plutellae bracovirus (CpBV) is a PDV symbiotic to an endoparasitoid wasp, C. plutellae, parasitizing young larvae of Plutella xylostella. An early expressed gene, CpBV-ELP1, plays an important role in the parasitism by suppressing host cellular immunity by its cytotoxic activity against hemocytes. This study aimed to test its oral toxicity against insect pest by expressing it in a recombinant tobacco plant. A recombinant CpBV-ELP1 protein was produced using a baculovirus expression system and secreted to cell culture medium. The cell cultured media were used to purify CpBV-ELP1 by a sequential array of purification steps: ammonium sulfate fractionation, size exclusion chromatography, and ion exchange chromatography. Purified rCpBV-ELP1 exhibited a significant cytotoxicity against Spodoptera exigua hemocytes. CpBV-ELP1 was highly toxic to the fifth instar larvae of S. exigua by injection to hemocoel. It also showed a significant oral toxicity to fifth instar larvae of S. exigua by a leaf-dipping assay. CpBV-ELP1 was cloned into pBI121 vector under CaMV 35S promoter with opaline synthase terminator. Resulting recombinant vector (pBI121-ELP1) was used to transform Agrobacterium tumefaciens LBA4404. The recombinant bacteria were then used to induce callus of a tobacco (Nicotiana tabacum Xanthi) leaves and subsequent generation (T1) plants were selected. T1 generation tobacco plants expressing CpBV-ELP1 gave significant insecticidal activities against S. exigua larvae. These results suggest that CpBV-ELP1 gene can be used to control insect pests by constructing transgenic crops.

• KSBB Journal
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• 제24권1호
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• pp.80-88
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• 2009

Human prostatic acid phosphatase (PAP), with comprehensive homology to glandular kallikrein, are representative serum biomarkers of prostate cancer. Dendritic cell (DC), which is the potent antigen-presenting cells(APC) in the immune system, can induce strong T cell responses against viruses, microbial pathogens, and tumors. Therefore, the immunization using DC loaded with tumor-associated antigens is a powerful method for inducing anti-tumor immunity. The CTP (Cytoplasmic Transduction Peptide) technology developed by Creagene which can transport attached bio-polymers like nucleic acids or proteins into the cell with high permeation efficiency. As the active forms of PAP can mediate apoptotic processing, we used multimer forms of PAP as an inactive form for antigen pulsing of DCs. In this study, multimeric forms of CTP-rhPAP was obtained according to the advanced purification process and subsequently confirmed by gel filtration chromatography, western blot and Dynamic Light Scattering. Therefore, CTP-conjugated PA multimers transduced into the cytoplasm were efficiently presented on the cell surface without any harm effect on cells via MHC class I molecules and result in induction of a large number of effector cell.

• Asian Pacific Journal of Cancer Prevention
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• 제13권8호
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• pp.4031-4036
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• 2012

Background: The negative signaling provided by interactions of the co-inhibitory molecule, programmed death-1 (PD-1), and its ligands, B7-H1 (PD-L1) and B7-DC (PD-L2), is a critical mechanism contributing to tumor evasion; blockade of this pathway has been proven to enhance cytotoxic activity and mediate antitumor therapy. Here we evaluated the anti-tumor efficacy of AAV-mediated delivery of the extracellular domain of murine PD-1 (sPD-1) to a tumor site. Material and Methods: An rAAV vector was constructed in which the expression of sPD-1, a known negative regulator of TCR signals, is driven by human cytomegalovirus immediate early promoter (CMV-P), using a triple plasmid transfection system. Tumor-bearing mice were then treated with the AAV/sPD1 construct and expression of sPD-1 in tumor tissues was determined by semi quantitative RT-PCR, and tumor weights and cytotoxic activity of splenocytes were measured. Results: Analysis of tumor homogenates revealed sPD-1 mRNA to be significantly overexpressed in rAAV/sPD-1 treated mice as compared with control levels. Its use for local gene therapy at the inoculation site of H22 hepatoma cells could inhibit tumor growth, also enhancing lysis of tumor cells by lymphocytes stimulated specifically with an antigen. In addition, PD-1 was also found expressed on the surfaces of activated CD8+ T cells. Conclusion: This study confirmed that expression of the soluble extracellular domain of PD-1 molecule could reduce tumor microenvironment inhibitory effects on T cells and enhance cytotoxicity. This suggests that it might be a potential target for development of therapies to augment T-cell responses in patients with malignancies.

• Korean Journal of Microbiology
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• 제53권1호
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• pp.1-8
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• 2017

Measles virus is a highly contagious, exanthematic virus, preventable by the use of an effective live-attenuated vaccine. However, measles virus remains endemic in many area of the world causing nearly 200,000 deaths per year and still a major cause of child mortality, mostly in developing countries. In March 2014, Republic of Korea was certified as a 'national measles elimination' by the WHO as a result of a high-quality case-based surveillance system and population immunity, which was achieved by a high vaccination rate (>95.0% since 1996). But, since the beginning of 2014, the Gyeonggi province has experienced a resurgence of measles cases. In this study, we investigated the characteristics of measles viruses isolated from confirmed measles in Gyeonggi province during January 1, 2014 ~ July 31, 2014, 60 isolates were obtained from 72 confirmed measles specimens. Genotypic distributions and genetic diversities of isolated measles virus were analyzed by sequencing of nucleoprotein (N) gene. 58 (96.7%) imported cases were identified. The predominant genotype was B3, which reflects the circulating measles virus in adjacent countries. The sequences of nucleoprotein (N) gene of isolated MeV were showed that the strains characterized showed the highest degree of identity (99%) with the Philippine related strains in 2013-2014. Therefore, infected traveler returning from the Philippines transmitted secondary infection in Korea.

• Journal of Microbiology and Biotechnology
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• 제26권6호
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• pp.1035-1045
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• 2016

To evaluate the effects of lactic acid bacteria (LAB) on Peyer's patch cells, mice were treated with a high dose of kanamycin to disturb the gut microbial environment. The overarching goal was to explore the potential of LAB for use as a dietary probiotic that buffers the negative consequences of antibiotic treatment. In vitro, LAB stimulated the production of immunoglobulin A (IgA) from isolated Peyer's patch cells. Inflammation-related genes (TNF-α, IL-1β, and IL-8) were up-regulated in Caco-2 cells stimulated with lipopolysaccharide (LPS), while tight-junction-related genes (ZO-1 and occludin) were down-regulated; the effects of LPS on inflammatory gene and tight-junction gene expression were reversed by treatment with LAB. Mice treated with a high dose of kanamycin showed increased serum IgE levels and decreases in serum IgA and fecal IgA levels; the number of Peyer's patch cells decreased with kanamycin treatment. However, subsequent LAB treatment was effective in reducing the serum IgE level and recovering the serum IgA and fecal IgA levels, as well as the number of Peyer's patch cells. In addition, ZO-1 and occludin mRNA levels were up-regulated in the ileum tissues of mice receiving LAB treatment. Lactic acid bacteria can enhance the intestinal immune system by improving the integrity of the intestinal barrier and increasing the production of IgA in Peyer's patches. Lactic acid bacteria should be considered a potential probiotic candidate for improving intestinal immunity, particularly in mitigating the negative consequences of antibiotic use.

• BMB Reports
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• 제55권2호
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• pp.81-86
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• 2022

Macrophages are a major cellular component of innate immunity and are mainly known to have phagocytic activity. In the tumor microenvironment (TME), they can be differentiated into tumor-associated macrophages (TAMs). As the most abundant immune cells in the TME, TAMs promote tumor progression by enhancing angiogenesis, suppressing T cells and increasing immunosuppressive cytokine production. N-myc downstream-regulated gene 2 (NDRG2) is a tumor suppressor gene, whose expression is down-regulated in various cancers. However, the effect of NDRG2 on the differentiation of macrophages into TAMs in breast cancer remains elusive. In this study, we investigated the effect of NDRG2 expression in breast cancer cells on the differentiation of macrophages into TAMs. Compared to tumor cell-conditioned medium (TCCM) from 4T1-mock cells, TCCM from NDRG2-over-expressing 4T1 mouse breast cancer cells did not significantly change the morphology of RAW 264.7 cells. However, TCCM from 4T1-NDRG2 cells reduced the mRNA levels of TAM-related genes, including MR1, IL-10, ARG1 and iNOS, in RAW 264.7 cells. In addition, TCCM from 4T1-NDRG2 cells reduced the expression of TAM-related surface markers, such as CD206, in peritoneal macrophages (PEM). The mRNA expression of TAM-related genes, including IL-10, YM1, FIZZ1, MR1, ARG1 and iNOS, was also downregulated by TCCM from 4T1-NDRG2 cells. Remarkably, TCCM from 4T1-NDRG2 cells reduced the expression of PD-L1 and Fra-1 as well as the production of GM-CSF, IL-10 and ROS, leading to the attenuation of T cell-inhibitory activity of PEM. These data showed that compared with TCCM from 4T1-mock cells, TCCM from 4T1-NDRG2 cells suppressed the TAM differentiation and activation. Collectively, these results suggest that NDRG2 expression in breast cancer may reduce the differentiation of macrophages into TAMs in the TME.

• Animal Bioscience
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• 제35권3호
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• pp.461-474
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• 2022

Objective: This experiment investigated the effects of supplementing vitamin D₃-fortified sow and progeny diets with 25(OH)D₃ on growth performance, carcass characteristics, immunity, and pork meat quality. Methods: The present study involved the assessment of supplementing the diet of sows and their progeny with or without 25 (OH)D₃ in a 2×2 factorial arrangement on the performance and production characteristics of wean-finish pigs. Forty-eight multiparous sows were assigned to a basal diet containing 2000 IU/kg vitamin D₃ and supplemented without (CON) or with (TRT) 50 ㎍/kg 25 (OH)D₃. At weaning, a total of 80 pigs each from CON and TRT sows were allocated to weaning and growing-finishing basal diets fortified with 2,500 and 1,750 IU/kg vitamin D₃ respectively and supplemented without or with 50 ㎍/kg 25(OH)D₃. Results: Sows fed 25(OH)D₃-supplemented diets improved pre-weaning growth rate of nursing piglets. A significant sow and pig weaning diet effect was observed for growth rate and feed efficiency (p<0.05) during days 1 to 42 post-weaning. Pigs consuming 25(OH)D₃-supplemented diets gained weight faster (p = 0.016), ate more (p = 0.044) and tended to convert feed to gain more efficiently (p = 0.088) than those fed CON diet between days 98 and 140 post-weaning. Supplemental 25(OH)D₃ improved water holding capacity and reduced drip loss of pork meat, increased serum 25(OH)D₃ level, produced higher interleukin-1 and lower interleukin-6 concentrations in blood circulation, downregulated myostatin (MSTN) and upregulated myogenic differentiation (MYOD) and myogenic factor 5 (MYF5) gene expressions (p<0.05). Conclusion: Supplementing vitamin D₃-fortified sow and wean-finish pig diets with 50 ㎍/kg 25(OH)D₃ significantly improved production performance suggesting their current dietary vitamin D₃ levels are insufficient. In fulfilling the total need for vitamin D, it is strongly recommended to add 50 ㎍/kg 25(OH)D₃ "on top" to practical vitamin D₃-fortified sow and wean-finish pig diets deployed under commercial conditions.

• Journal of Veterinary Science
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• 제24권5호
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• pp.73.1-73.16
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• 2023

Background: Highly pathogenic avian influenza virus (HPAIV) is considered a global threat to both human health and the poultry industry. MicroRNAs (miRNA) can modulate the immune system by affecting gene expression patterns in HPAIV-infected chickens. Objectives: To gain further insights into the role of miRNAs in immune responses against H5N1 infection, as well as the development of strategies for breeding disease-resistant chickens, we characterized miRNA expression patterns in tracheal tissues from H5N1-infected Ri chickens. Methods: miRNAs expression was analyzed from two H5N1-infected Ri chicken lines using small RNA sequencing. The target genes of differentially expressed (DE) miRNAs were predicted using miRDB. Gene Ontology and Kyoto Encyclopedia of Genes and Genomes analysis were then conducted. Furthermore, using quantitative real-time polymerase chain reaction, we validated the expression levels of DE miRNAs (miR-22-3p, miR-146b-3p, miR27b-3p, miR-128-3p, miR-2188-5p, miR-451, miR-205a, miR-203a, miR-21-3p, and miR-200a3p) from all comparisons and their immune-related target genes. Results: A total of 53 miRNAs were significantly expressed in the infection samples of the resistant compared to the susceptible line. Network analyses between the DE miRNAs and target genes revealed that DE miRNAs may regulate the expression of target genes involved in the transforming growth factor-beta, mitogen-activated protein kinase, and Toll-like receptor signaling pathways, all of which are related to influenza A virus progression. Conclusions: Collectively, our results provided novel insights into the miRNA expression patterns of tracheal tissues from H5N1-infected Ri chickens. More importantly, our findings offer insights into the relationship between miRNA and immune-related target genes and the role of miRNA in HPAIV infections in chickens.

• Journal of Plant Biotechnology
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• 제43권3호
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• pp.347-358
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• 2016

Brassinosteroid (BR), a plant steroid hormone, plays key roles in numerous growth and developmental processes as well as tolerance to both abiotic and biotic stress. To understand the biological networks involved in BR-mediated signaling pathways and stress tolerance, we performed comparative genome-wide transcriptome analysis of a constitutively activated BR bes1-D mutant with an Agilent Arabidopsis $4{\times}44K$ oligo chip. As a result, we newly identified 1,091 (562 up-regulated and 529 down-regulated) significant differentially expressed genes (DEGs). The combination of GO enrichment and protein network analysis revealed that stress-related processes, such as metabolism, development, abiotic/biotic stress, immunity, and defense, were critically linked to BR signaling pathways. Among the identified gene sets, we confirmed more than a 6-fold up-regulation of NB-ARC and FLS2 in bes1-D plants. However, some genes, including TIR1, TSA1 and OCP3, were down-regulated. Consistently, BR-activated plants showed higher tolerance to drought stress and pathogen infection compared to wild-type controls. In this study, we newly developed a useful, comprehensive method for large-scale identification of critical network and gene sets with global transcriptome analysis using a microarray. This study also showed that gain of function in the bes1-D gene can regulate the adaptive response of plants to various stressful conditions.

• Journal of the Korean Society of Food Science and Nutrition
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• 제46권3호
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• pp.306-313
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• 2017

Purple bamboo salt (PuBS) is commonly used as a medicinal food in Korea and has beneficial potentials such as antioxidant and anti-inflammatory effects. Rubus coreanus is called Bokbunja, which is used as a traditional medicine for treating asthma, impotence, and allergic diseases in Korea. The aim of present study was to investigate the immunostimulatory effect of PuBS composed with Rubus coreanus (PuBS-R). We performed comparative analysis between PuBS and PuBS-R in Raw264.7 cells, which is a mouse macrophage cell line, and peritoneal macrophages isolated primarily from the mouse peritoneal cavity. We evaluated cytotoxicity and the immune cytokine response in PuBS- and PuBS-R-treated cells. Both PuBS and PuBS-R did not have any cytotoxicity in Raw264.7 cells up to $500{\mu}g/mL$. Gene and protein levels of immune cytokines such as tumor necrosis $factor-{\alpha}$ ($TNF-{\alpha}$), $interferon-{\gamma}$ ($IFN-{\gamma}$), interleukin (IL)-10, and IL-12 were significantly elevated by PuBS-R more than PuBS in Raw264.7 cells. Moreover, we evaluated the immunostimulatory effects of PuBS-R on mouse primary peritoneal macrophages. Protein levels of inducible nitric oxide synthase, $TNF-{\alpha}$, $IFN-{\gamma}$, IL-10, and IL-12 were significantly higher in PuBS-R-treated peritoneal macrophages than PuBS-treated peritoneal macrophages. These results suggest the potential immunostimulatory effect of PuBS-R for immunity against harmful infection.