• Journal of Veterinary Science
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• v.24 no.5
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• pp.72.1-72.16
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• 2023

Background: Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) on the surface of Streptococcus dysgalactiae, coded with gapC, is a glycolytic enzyme that was reported to be a moonlighting protein and virulence factor. Objective: This study assessed GAPDH as a potential immunization candidate protein to prevent streptococcus infections. Methods: Mice were vaccinated subcutaneously with recombinant GAPDH and challenged with S. dysgalactiae in vivo. They were then evaluated using histological methods. rGAPDH of mouse bone marrow-derived dendritic cells (BMDCs) was evaluated using immunoblotting, reverse transcription quantitative polymerase chain reaction, and enzyme-linked immunosorbent assay methods. Results: Vaccination with rGAPDH improved the survival rates and decreased the bacterial burdens in the mammary glands compared to the control group. The mechanism by which rGAPDH vaccination protects against S. dysgalactiae was investigated. In vitro experiments showed that rGAPDH boosted the generation of interleukin-10 and tumor necrosis factor-α. Treatment of BMDCs with TAK-242, a toll-like receptor 4 inhibitor, or C29, a toll-like receptor 2 inhibitor, reduced cytokines substantially, suggesting that rGAPDH may be a potential ligand for both TLR2 and TLR4. Subsequent investigations showed that rGAPDH may activate the phosphorylation of MAPKs and nuclear factor-κB. Conclusions: GAPDH is a promising immunization candidate protein for targeting virulence and enhancing immune-mediated protection. Further investigations are warranted to understand the mechanisms underlying the activation of BMDCs by rGAPDH in a TLR2- and TLR4-dependent manner and the regulation of inflammatory cytokines contributing to mastitis pathogenesis.

• Journal of fish pathology
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• v.37 no.1
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• pp.1-8
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• 2024

The advantages of replicon vectors of RNA viruses include a high ability to stimulate innate immunity and exponential amplification of target mRNA leading to high expression of foreign antigens. The present study aimed to construct a DNA-layered nervous necrosis virus (NNV) replicon vector system in which the capsid protein gene was replaced with a foreign antigen gene and to compare the efficiency of foreign antigen expression between the conventional DNA vaccine vector and the present replicon vector. We presented the first report of a nodavirus DNA replicon-based foreign antigen expression system. Instead of a two-vector system, we devised a one-vector system containing both an NNV RNA-dependent RNA polymerase cassette and a foreign antigen-expressing cassette. This single-vector approach circumvents the issue of low foreign protein expression associated with the low co-transfection efficiency of a two-vector system. Cells transfected with a vector harboring hammerhead ribozyme-fused RNA1 and RNA2 (with the capsid gene ORF replaced with VHSV glycoprotein ORF) exhibited significantly higher transcription of the VHSV glycoprotein gene compared to cells transfected with either a vector without hammerhead ribozyme or a conventional DNA vaccine vector expressing the VHSV glycoprotein. Furthermore, the transcription level of the VHSV glycoprotein in cells transfected with a vector harboring hammerhead ribozyme-fused RNA1 and RNA2 showed a significant increase over time. These results suggest that NNV genome-based DNA replicon vectors have the potential to induce stronger and longer expression of target antigens compared to conventional DNA vaccine vectors.

• IMMUNE NETWORK
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• v.21 no.6
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• pp.42.1-42.20
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• 2021

Eosinophils play critical roles in the maintenance of homeostasis in innate and adaptive immunity. Although primarily known for their roles in parasitic infections and the development of Th2 cell responses, eosinophils also play complex roles in other immune responses ranging from anti-inflammation to defense against viral and bacterial infections. However, the contributions of pattern recognition receptors in general, and NOD-like receptors (NLRs) in particular, to eosinophil involvement in these immune responses remain relatively underappreciated. Our in vivo studies demonstrated that NLRC4 deficient mice had a decreased number of eosinophils and impaired Th2 responses after induction of an allergic airway disease model. Our in vitro data, utilizing human eosinophilic EoL-1 cells, suggested that TLR2 induction markedly induced pro-inflammatory responses and inflammasome forming NLRC4 and NLRP3. Moreover, activation by their specific ligands resulted in caspase-1 cleavage and mature IL-1β secretion. Interestingly, Th2 responses such as secretion of IL-5 and IL-13 decreased after transfection of EoL-1 cells with short interfering RNAs targeting human NLRC4. Specific induction of NLRC4 with PAM3CSK4 and flagellin upregulated the expression of IL-5 receptor and expression of Fc epsilon receptors (FcεR1α, FcεR2). Strikingly, activation of the NLRC4 inflammasome also promoted expression of the costimulatory receptor CD80 as well as expression of immunoregulatory receptors PD-L1 and Siglec-8. Concomitant with NLRC4 upregulation, we found an increase in expression and activation of matrix metalloproteinase (MMP)-9, but not MMP-2. Collectively, our results present new potential roles of NLRC4 in mediating a variety of eosinopilic functions.

• IMMUNE NETWORK
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• v.20 no.1
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• pp.5.1-5.15
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• 2020

The γδ T cells are unconventional lymphocytes that function in both innate and adaptive immune responses against various intracellular and infectious stresses. The γδ T cells can be exploited as cancer-killing effector cells since γδ TCRs recognize MHC-like molecules and growth factor receptors that are upregulated in cancer cells, and γδ T cells can differentiate into cytotoxic effector cells. However, γδ T cells may also promote tumor progression by secreting IL-17 or other cytokines. Therefore, it is essential to understand how the differentiation and homeostasis of γδ T cells are regulated and whether distinct γδ T cell subsets have different functions. Human γδ T cells are classified into Vδ2 and non-Vδ2 γδ T cells. The majority of Vδ2 γδ T cells are Vγ9δ2 T cells that recognize pyrophosphorylated isoprenoids generated by the dysregulated mevalonate pathway. In contrast, Vδ1 T cells expand from initially diverse TCR repertoire in patients with infectious diseases and cancers. The ligands of Vδ1 T cells are diverse and include the growth factor receptors such as endothelial protein C receptor. Both Vδ1 and Vδ2 γδ T cells are implicated to have immunotherapeutic potentials for cancers, but the detailed elucidation of the distinct characteristics of 2 populations will be required to enhance the immunotherapeutic potential of γδ T cells. Here, we summarize recent progress regarding cancer immunology of human γδ T cells, including their development, heterogeneity, and plasticity, the putative mechanisms underlying ligand recognition and activation, and their dual effects on tumor progression in the tumor microenvironment.

• Journal of Integrative Natural Science
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• v.17 no.2
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• pp.59-65
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• 2024

Dendritic cells play a very important role in the immune response as antigen-presenting cells that are critical for initiating both innate and acquired immunity. They recognize, process and present foreign antigens to other key immune cells to trigger and regulate the immune response. The ability to activate these dendritic cells can be used as a treatment for various immune diseases. Maqui berry has been reported to have anticancer, antibacterial and anti-inflammatory properties. However, its effect on the activity of dendritic cells has not been studied. In this study, we investigated the efficacy of maqui berry extract in modulating dendritic cell activity. Treatment of dendritic cells with maqui berry extract induced the costimulatory molecules CD80, CD86, and MHC class I and II in a concentration-dependent manner. Furthermore, the antigen-presenting capacity of dendritic cells was inhibited, which confirms their ability to present antigens, and the production of Interleukin (IL)-12, which is important for dendritic cell activity, was increased. These results indicated that Maqui berry extract activates dendritic cells maturation by inducing the production of co-stimulatory molecules and IL-12. These results suggest that maqui berry extract may act as an effective adjuvant to enhance dendritic cell-based immune responses.

• Journal of Microbiology and Biotechnology
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• v.34 no.2
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• pp.233-239
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• 2024

N6-methyladenosine (m6A) RNA methylation has recently emerged as a significant co-transcriptional modification involved in regulating various RNA functions. It plays a vital function in numerous biological processes. Enzymes referred to as m6A methyltransferases, such as the methyltransferase-like (METTL) 3-METTL14-Wilms tumor 1 (WT1)-associated protein (WTAP) complex, are responsible for adding m6A modifications, while m6A demethylases, including fat mass and obesity-associated protein (FTO) and alkB homolog 5 (ALKBH5), can remove m6A methylation. The functions of m6A-methylated RNA are regulated through the recognition and interaction of m6A reader proteins. Recent research has shown that m6A methylation takes place at multiple sites within hepatitis B virus (HBV) RNAs, and the location of these modifications can differentially impact the HBV infection. The addition of m6A modifications to HBV RNA can influence its stability and translation, thereby affecting viral replication and pathogenesis. Furthermore, HBV infection can also alter the m6A modification pattern of host RNA, indicating the virus's ability to manipulate host cellular processes, including m6A modification. This manipulation aids in establishing chronic infection, promoting liver disease, and contributing to pathogenesis. A comprehensive understanding of the functional roles of m6A modification during HBV infection is crucial for developing innovative approaches to combat HBV-mediated liver disease. In this review, we explore the functions of m6A modification in HBV replication and its impact on the development of liver disease.

• Journal of Rhinology
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• v.59 no.1
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• pp.49-58
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• 2021

Background: Nasal polyps in the nasal cavity and mucous discharge inside the maxillary sinus exhibit compressive stress on the nasal mucosal epithelium. However, there have been only a few studies on how compressive stress impacts the human nasal mucosal epithelium. Methodology: We investigated the effect of compressive stress on collective migration, junctional proteins, transepithelial electrical resistance, epithelial permeability, and gene expression in well-differentiated normal human nasal epithelial (NHNE) cells and human nasal polyp epithelial (HNPE) cells. Results: NHNE cells barely showed collective migration at compressive stress up to 150 mmH₂0. However, HNPE cells showed much greater degree of collective migration at a lower compressive stress of 100 mmH₂0. The cell migration of HNPE cells subjected to 100 mmH₂O compression was significantly decreased at day 3 and was recovered to the status prior to the compressive stress by day 7, indicating that HNPE cells are relatively more sensitive to mechanical pressure than NHNE cells. Compressive stress also increased transepithelial electrical resistance and decreased epithelial permeability, indicating that the compressive stress disturbed the structural organization rather than physical interactions between cells. In addition, we found that compressive stress induced gene expressions relevant to airway inflammation and tissue remodelling in HNPE cells. Conclusion: Taken together, these findings demonstrate that compressive stress on nasal polyp epithelium is capable of inducing collective migration and induce increased expression of genes related to airway inflammation, innate immunity, and polyp remodelling, even in the absence of inflammatory mediators.

• Pediatric Infection and Vaccine
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• v.13 no.1
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• pp.63-70
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• 2006

Purpose : We hypothesized that mannose binding lectin gene(MBL2), a key molecule of innate immunity, may contirbute to the development and the outcome of respiratory syncytial virus(RSV) disease in early childhood. This study was performed to investigate the genetic basis of polymorphisms and haplotypes of MBL2 for RSV disease severity in Korean children. Methods : Cases with severe RSV diseases are 99 children with severe RSV lower respiratory tract infections, who were admitted to the Seoul National University Children's Hospital through 1993~2000. The control subjects consisted of 224 anonymous healthy Korean blood donors. The frequency of promoter variant(-221, X/Y) and structural variant(codon 54) were compared between the case patient group and the control subject group. Results : The mean age of patients was 11.8 months; 49% were <6 months, 39% were 6-24 months and 12% were >24 months. In the cohort of cases of severe RSV diseases, the genotypic frequencies of structural variant in codon 54 were 61% for AA, 34% for AB, and 5% for BB. Those of the promoter X/Y variant were 85% for YY and 15% for XY. There were no significant differences in overall distribution of both structural and promoter variants between the cases and the control subjects. We did not observe statistical difference in the haplotypic frequencies of MBL2. Conclusion : Common variants of MBL2 gene most likely do not contribute to the risk for severe RSV diseases in Korean children. Further genetic association studies should be conducted in a larger propsectively recruited cohort of children with RSV infection.

• Journal of Life Science
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• v.28 no.11
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• pp.1301-1315
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• 2018

We purified an antimicrobial peptide from the acidified hemocyte extract of Mytilus coruscus by $C_{18}$ reversed-phase high-performance liquid chromatography (RP-HPLC). The peptide was 4041.866 Da based on matrix-assisted laser desorption ionization time-of-flight mass spectrophotometer (MALDI-TOF/MS) and the 25 amino acids of the N-terminus sequence were identified. Comparison of this sequence of the purified peptide with the N-terminus sequences of other antimicrobial peptides revealed 100% identity with the mytilin B precursor of Mytilus coruscus. We also identified a 312 bp open-reading frame (ORF) encoding 103 amino acids based on the obtained amino acid residues. The nucleotide sequence of this ORF and the amino acid sequence also revealed 100% identity with the mytilin B precursor of Mytilus coruscus. We synthesized two antimicrobial peptides with an alanine residue in the C-terminus, and designated them mytilin B1 and B2. These two antimicrobial peptides showed antimicrobial activity against gram-positive bacteria, including Bacillus cereus and Streptococcus parauberis (minimal effective concentration, MECs $41.6-89.7{\mu}g/ml$), gram-negative bacteria, including Enterobacter cloacae, Escherichia coli, Klebsiella pneumoniae, Proteus mirabilis, Providencia stuartii, Pseudomonas aeruginosa, and Vibrio ichthyoenteri (MECs $7.4-39.5{\mu}g/ml$), and the fungus Candida albicans (MECs $26.0-31.8{\mu}g/ml$). This antimicrobial activity was stable under heat and salt conditions. Furthermore, the peptides did not exhibit significant hemolytic activity or cytotoxic effects. These results suggest that mytilin B could be applied as alternative antibiotic agent, and they add to the understanding of the innate immunity of hard-shelled mussels.

• Journal of Life Science
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• v.28 no.9
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• pp.1076-1080
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• 2018

Carrageenan (CGN) has been used as a safe food additive for several decades. CGN has also been widely used to induce inflammation in various animal models. Likewise, degraded CGN (dCGN), which is produced by subjecting CGN to acid hydrolysis, also induces inflammation and does so more effectively than CGN. One of the most important characteristics of an immunological adjuvant is its ability to activate innate immunity. The immune-adjuvant effects of CGN and dCGN have not yet been studied in detail. The purpose of this study was to evaluate the immunological adjuvant activities of both CGN and dCGN, which was done by comparing the levels of an ovalbumin (OVA)-specific antibody after treatment with OVA in the absence or presence of CGN or dCGN in plasma from immunized mice. CGN and dCGN showed similar levels of adjuvant activity, as evidenced by increased antibody titer. Specifically, both CGN and dCGN significantly increased the levels of OVA-specific IgG, IgG1, and IgG2a antibodies in the plasma as compared with OVA alone (the control). However, compared to the positive control (Freund's adjuvant), both CGN and dCGN caused greater increases in IgG1 than in IgG2a. These results suggest that CGN and dCGN have similar adjuvant activities and produce more IgG1 antibodies than IgG2a.