• Genes and Genomics
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• v.40 no.11
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• pp.1225-1235
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• 2018

Hypoxia seriously affects the innate immune system of fish. However, the roles of suppressor of cytokine signaling (SOCS), pivotal anti-inflammatory genes, in response to hypoxia/reoxygenation remain largely unexplored. The primary objective of this study was to elucidate the function of SOCS genes under acute hypoxia and reoxygenation in pufferfish (Takifugu fasciatus). In the present study, SOCS1, 2 and 3 were identified in T. fasciatus referred to as TfSOCS1, 2 and 3. Then, qRT-PCR and western blot analysis were employed to assess their expressions at both the mRNA and protein levels. Tissue distribution demonstrated that the three SOCS genes were predominantly distributed in gill, brain and liver. Under hypoxia challenge ($1.63{\pm}0.2mg/L$ DO for 2, 4, 6 and 8 h), the expressions of TfSOCS1 and 3 in brain and liver at the mRNA and protein levels were significantly decreased, while their expressions showed an opposite trend in gill. Different from the expressions of TfSOCS1 and 3, the expression of TfSOCS2 was inhibited in gill, along with its increased expression in brain and liver. After normoxic recovery ($7.0{\pm}0.3mg/L$ of DO for 4 and 12 h), most of TfSOCS genes were significantly altered at R4 (reoxygenation for 4 h) and returned to the normal level at R12 (reoxygenation for 12 h). SOCS genes played vital roles in response to hypoxia/reoxygenation challenge. Our findings greatly strengthened the relation between innate immune and hypoxia stress in T. fasciatus.

• Nutrition Research and Practice
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• v.13 no.2
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• pp.95-104
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• 2019

BACKGROUND/OBJECTIVES: Inflammatory Bowel Disease (IBD) has rapidly escalated in Asia (including Korea) due to increasing westernized diet patterns subsequent to industrialization. Factors associated with endoplasmic reticulum (ER) stress are demonstrated to be one of the major causes of IBD. This study was conducted to investigate the effect of Lycium barbarum (L. barbarum) on ER stress. MATERIALS/METHODS: Mouse embryonic fibroblast (MEF) cell line and polarized Caco-2 human intestinal epithelial cells were treated with crude extract of the L. chinense fruit (LF). Paracellular permeability was measured to examine the effect of tight junction (TJ) integrity. The regulatory pathways of ER stress were evaluated in MEF knockout (KO) cell lines by qPCR for interleukin (IL) 6, IL8 and XBP1 spliced form (XBP1s). Immunoglobulin binding protein (BiP), XBP1s and CCAAT/enhancer-binding homologous protein (CHOP) expressions were measured by RT-PCR. Scanning Ion Conductance Microscopy (SICM) at high resolution was applied to observe morphological changes after treatments. RESULTS: Exposure to LF extract strengthened the TJ, both in the presence and absence of inflammation. In polarized Caco-2 pretreated with LF, induction in the expression of proinflammatory marker IL8 was not significant, whereas ER stress marker XBP1s expression was significantly increased. In wild type (wt) MEF cells, IL6, CHOP and XBP1 spliced form were dose-dependently induced when exposed to $12.5-50{\mu}g/mL$ extract. However, absence of XBP1 or $IRE1{\alpha}$ in MEF cells abolished this effect. CONCLUSION: Results of this study show that LF treatment enhances the barrier function and reduces inflammation and ER stress in an $IRE1{\alpha}$-XBP1-dependent manner. These results suggest the preventive effect of LF on healthy intestine, and the possibility of reducing the degree of inflammatory symptoms in IBD patients.

• Asian-Australasian Journal of Animal Sciences
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• v.32 no.2
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• pp.170-175
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• 2019

Objective: Uncoupling protein 3 gene (UCP3) is a candidate gene associated with the meat quality of pigs. The aim of this study was to explore the regulation mechanism of UCP3 expression and provide a theoretical basis for the research of the function of porcine UCP3 gene in meat quality. Methods: Bisulfite sequencing polymerase chain reaction (PCR) and quantitative real-time PCR (Q-PCR) were used to analyze the methylation of UCP3 5′-flanking region and UCP3 mRNA expression in the adipose tissue or skeletal muscle of three pig breeds at different ages (1, 90, 210-day-old Putian Black pig; 90-day-old Duroc; and 90-day-old Dupu). Results: Results showed that two cytosine-guanine dinucleotide (CpG) islands are present in the promoter region of porcine UCP3 gene. The second CpG island located in the core promoter region contained 9 CpG sites. The methylation level of CpG island 2 was lower in the adipose tissue and skeletal muscle of 90-day-old Putian Black pigs compared with 1-day-old and 210-day-old Putian Black pigs, and the difference also existed in the skeletal muscle among the three 90-day-old pig breeds. Furthermore, the obvious changing difference of UCP3 mRNA expression was observed in the skeletal muscle of different groups. However, the difference of methylation status and expression level of UCP3 gene was not significant in the adipose tissue. Conclusion: Our data indicate that UCP3 mRNA expression level was associated with the methylation status of UCP3 promoter in the skeletal muscle of pigs.

• Journal of Ginseng Research
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• v.43 no.4
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• pp.666-675
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• 2019

Background: Korean Red Ginseng (KRG) has been widely used as an herbal medicine to normalize and strengthen body functions. Although many researchers have focused on the biological effects of KRG, more studies on the action mechanism of red ginseng are still needed. Previously, we investigated the proteomic changes of the rat spleen while searching for molecular signatures and the action mechanism of KRG. The proteomic analysis revealed that differentially expressed proteins (DEPs) were involved in the increased immune response and phagocytosis. The aim of this study was to evaluate the biological activities of KRG, especially the immune-enhancing response of KRG. Methods: Rats were divided into 4 groups: 0 (control group), 500, 1000, and 2000 mg/kg administration of KRG powder for 6 weeks, respectively. Isobaric tags for relative and absolute quantitation was performed with Q-Exactive LC-MS/MS to compare associated proteins between the groups. The putative DEPs were identified by a current UniProt rat protein database search and by the Gene Ontology annotations. Results: The DEPs appear to increase the innate and acquired immunity as well as immune cell movement. These results suggest that KRG can stimulate immune responses. This analysis refined our targets of interest to include the potential functions of KRG. Furthermore, we validated the potential molecular targets of the functions, representatively LCN2, CRAMP, and HLA-DQB1, by Western blotting. Conclusion: These results may provide molecular signature candidates to elucidate the mechanisms of the immune response by KRG. Here, we demonstrate a strategy of tissue proteomics for the discovery of the molecular function of KRG.

• Journal of Animal Reproduction and Biotechnology
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• v.34 no.1
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• pp.2-9
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• 2019

Because mesenchymal stem cells (MSCs) maintain distinct capacities with respect to self-renewal, differentiation ability and immunomodulatory function, they have been highly considered as the therapeutic agents for cell-based clinical application. Of particular, differentiation condition alters characteristics of MSCs, including cellular morphology, expression of gene/protein and cell surface molecule, immunological property and apoptosis. However, the previous results for differentiation-related apoptosis in MSCs have still remained controversial due to varied outcomes. Therefore, the present study aimed to disclose periodical alterations of pro- and anti-apoptosis in MSCs under differentiation inductions. The human dental pulp-derived MSCs (DP-MSCs) were differentiated into adipocytes and osteoblasts during early (1 week), middle (2 weeks) and late (3 weeks) stages, and were investigated on their apoptosis-related changes by Annexin V assay, qRT-PCR and western blotting. The ratio of apoptotic cell population was significantly (p < 0.05) elevated during the early to middle stages of differentiations but recovered up to the similar level of undifferentiated state at the late stage of differentiation. In the expression of mRNA and protein, whereas expressions of pro-apoptosis-related makers (BAX and BAK) were not altered in any kind and duration of differentiation inductions, anti-apoptosis marker (BCL2) was significantly (p < 0.05) elevated even at the early stage of differentiations. The recovery of apoptotic cell population at the late stage of differentiation is expected to be associated with the response by elevation of anti-apoptotic molecules. The present study may contribute on understanding for cellular mechanism in differentiation of MSCs and provide background data in clinical application of MSCs in the animal biotechnology to develop effective and safe therapeutic strategy.

• Yonsei Medical Journal
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• v.59 no.9
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• pp.1064-1071
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• 2018

Purpose: To explore the influence of S100 calcium binding protein A4 (S100A4) knockout (KO) on methionine-choline-deficient (MCD) diet-induced non-alcoholic fatty liver disease (NAFLD) in mice. Materials and Methods: S100A4 KO mice (n=20) and their wild-type (WT) counterparts (n=20) were randomly divided into KO/MCD, Ko/methionine-choline-sufficient (MCS), WT/MCD, and WT/MCS groups. After 8 weeks of feeding, blood lipid and liver function-related indexes were measured. HE, Oil Red O, and Masson stainings were used to observe the changes of liver histopathology. Additionally, expressions of S100A4 and proinflammatory and profibrogenic cytokines were detected by qRT-PCR and Western blot, while hepatocyte apoptosis was revealed by TUNEL staining. Results: Serum levels of aminotransferase, aspartate aminotransferase, triglyceride, and total cholesterol in mice were increased after 8-week MCD feeding, and hepatocytes performed varying balloon-like changes with increased inflammatory cell infiltration and collagen fibers; however, these effects were improved in mice of KO/MCD group. Meanwhile, total NAFLD activity scores and fibrosis were lower compared to WT+MCD group. Compared to WT/MCS group, S100A4 expression in liver tissue of WT/MCD group was enhanced. The expression of proinflammatory ($TGF-{\alpha}$, $IL-1{\beta}$, IL-6) and profibrogenic cytokines ($TGF-{\beta}1$, COL1A1, ${\alpha}-SMA$) in MCD-induced NAFLD mice were increased, as well as apoptotic index (AI). For MCD group, the expressions of proinflammatory and profibrogenic cytokines and AI in KO mice were lower than those of WT mice. Conclusion: S100A4 was detected to be upregulated in NAFLD, while S100A4 KO alleviated liver fibrosis and inflammation, in addition to inhibiting hepatocyte apoptosis.

• BMB Reports
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• v.54 no.9
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• pp.476-481
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• 2021

Liver receptor homolog-1 (LRH-1) has emerged as a regulator of hepatic glucose, bile acid, and mitochondrial metabolism. However, the functional mechanism underlying the effect of LRH-1 on lipid mobilization has not been addressed. This study investigated the regulatory function of LRH-1 in lipid metabolism in maintaining a normal liver physiological state during fasting. The Lrh-1^f/f and LRH-1 liver-specific knockout (Lrh-1^LKO) mice were either fed or fasted for 24 h, and the liver and serum were isolated. The livers were used for qPCR, western blot, and histological analysis. Primary hepatocytes were isolated for immunocytochemistry assessments of lipids. During fasting, the Lrh-1^LKO mice showed increased accumulation of triglycerides in the liver compared to that in Lrh-1^f/f mice. Interestingly, in the Lrh-1^LKO liver, decreases in perilipin 5 (PLIN5) expression and genes involved in β-oxidation were observed. In addition, the LRH-1 agonist dialauroylphosphatidylcholine also enhanced PLIN5 expression in human cultured HepG2 cells. To identify new target genes of LRH-1, these findings directed us to analyze the Plin5 promoter sequence, which revealed -1620/-1614 to be a putative binding site for LRH-1. This was confirmed by promoter activity and chromatin immunoprecipitation assays. Additionally, fasted Lrh-1^f/f primary hepatocytes showed increased co-localization of PLIN5 in lipid droplets (LDs) compared to that in fasted Lrh-1^LKO primary hepatocytes. Overall, these findings suggest that PLIN5 might be a novel target of LRH-1 to mobilize LDs, protect the liver from lipid overload, and manage the cellular needs during fasting.

• Journal of The Korean Society of Integrative Medicine
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• v.9 no.1
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• pp.163-171
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• 2021

Purpose : The purpose of this study was to examine the anti-inflammatory effects of Sparassis crispa (SC). SC is a well-known traditional herbal remedy and its mushroom is used for treatment of inflammation. Many diseases that are increasing recently have characteristics of inflammatory diseases. Researchers are finding bioactive substances from natural products that can promote treatment and prevention of inflammation. We investigated the effect of water extracted from SC on the expression of effector genes involved in the function of RAW 264.7 cells. Methods : Effects of RAW 264.7 cells on cell viability, antioxidation, and mRNA expression were examined using water extracts from SC. A 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay was performed to determine the effect of water extracts from SC on cell viability in RAW 264.7 cells. Inflammation of RAW 264.7 cells induced by lipopolysaccharide (LPS) treatment and expression levels of inflammatory cytokine TNF-α, iNOS and IL-1β gene were analyzed using quantitative reverse transcription PCR (qRT-PCR) analysis. Results : The MTS assay was performed on RAW 264.7 cells after treatment with various concentrations of water extracts of SC. Treatment of RAW 264.7 cells with water extracts from SC and LPS at a concentration of 0.125, 0.5 mg/㎖ for twenty four hours promoted mRNA expression of TNF-α, iNOS and IL-1β. Conclusion : MTS assay was applied to RAW 264.7 cells after various concentrations of water extracts of SC. Through experimental demonstration of anti-oxidant and anti-inflammatory effects of water extracts from SC, we suggest that SC is a valuable material for the prevention and treatment of various inflammatory diseases.

• Journal of Microbiology and Biotechnology
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• v.31 no.6
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• pp.794-802
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• 2021

In this study we investigated the role and mechanism of cardamonin on IL-1β induced injury in OA. CHON-001 cells were treated with cardamonin and IL-1β and transfected with silencing nuclear factor erythroid 2-related factor 2 (siNrf2). Cell viability was detected by Cell Counting Kit-8 assay and flow cytometer assay was utilized for cell apoptosis assessment. IL-6, IL-8, TNF-α and Nrf2 mRNA expression was tested by qRT-PCR. Western blot was employed to evaluate MMP-3, MMP-13, Collagen II, Nrf2, NQO-1, NLRP3, Caspase 1 and apoptosis-associated speck-like protein containing a caspase-1 recruitment domain (ASC) protein levels. In CHON-001 cells, IL-1β suppressed cell viability and Collagen II level while promoting cell apoptosis and expression of pro-inflammatory cytokines (IL-6, IL-8, TNF-α), MMPs (MMP-3, MMP-13), NQO-1, and NLRP3 inflammasome (NLRP3, Caspase 1 and ASC), with no significant influence on Nrf2. Cardamonin reversed the effect of IL-1β on cell viability, cell apoptosis, pro-inflammatory cytokines, MMPs, Collagen II, and NLRP3 inflammasome levels. In addition, cardamonin advanced Nrf2 and NQO-1 expression of CHON-001 cells. SiNrf2 reversed the function of cardamonin on IL-1β-induced cell apoptosis and expression of pro-inflammatory cytokines, Nrf2, NQO-1, and NLRP3 inflammasome in chondrocytes. Taken together Cardamonin inhibited IL-1β induced injury by inhibition of NLRP3 inflammasome via activating Nrf2/NQO1 signaling pathway in chondrocyte.

• The Korea Journal of Herbology
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• v.36 no.1
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• pp.97-102
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• 2021

Objectives : Virus infection through the respiratory tract causes various inflammatory diseases such as pneumonia, cystic fibrosis, and obstructive pulmonary disease, causing enormous social damage. Therefore, it is very important to develop a treatment and prevention of infectious diseases. In this study, we investigated the effect of water extracts of Liriope muscari (WELM), known to improve lung function, on the inflammatory response of lung carcinoma cell line A549 cells induced by the viral double stranded RNA mimetic Polyinosinic:polycytidylic acid (Poly I:C). Methods : The cell viability by WELM treatment was analyzed using MTS assay in A549 cells. After inducing an inflammatory response to WELM-treated A549 cells with Poly I:C, the degree of apoptosis was confirmed through bright field microscopy. Interferon beta (IFN-β) mRNA expression level in A549 cells was analyzed by quantitative reverse transcription PCR (qRT-PCR). Results : WELM treatment has no significant effect on cell viability of A549 cells. We confirmed that pre-treatment of WELM effectively reduces the Poly I:C-induced apoptotic cell death in A549 cells. In addition, it was confirmed that the mRNA expression level of IFN-β, a pro-inflammatory cytokine increased by Poly I:C treatment, was significantly suppressed by WELM treatment in A549 cells. Conclusions : These results provide the evidence that WELM is effective at inhibiting inflammation on respiratory viral infections and suggest that Liriope muscari might be a valuable natural substance in the prevention and treatment of infectious diseases.