• Tuberculosis and Respiratory Diseases
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• v.62 no.4
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• pp.299-307
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• 2007

Background: High mobility group box 1 (HMGB1) is a novel, late mediator of inflammation. This study compared the pro-inflammatory effects of LPS and HMGB1. The transcriptional factors that play an important role in mediating the HMGB1-induced stimulation of IL-8 were also evaluated. Methods: RAW264.7 cells were stimulated with either LPS (100 ng/ml) or HMGB1 (500 ng/ml). The $TNF-{\alpha}$, MIP-2 and $IL-1{\beta}$ levels in the supernatant were evaluated by ELISA at 0, 2, 4, 8, 12 and 24h after stimulation. An acute lung injury was induced by an injection of LPS (5 mg/kg) or HMGB1 (2.5 mg/kg) into the peritoneum of the Balb/c mice. The lung cytokines and MPO activity were measured at 4h (for LPS) or 24h (for HMGB1) after the injection. The transcriptional factor binding sites for NF-IL6, $NF-{\kappa}B$ and AP-1 in the IL-8 promoter region were artificially mutated. Each mutant was ligated with pIL-6luc and transfected into the RAW264.7 cells. One hour after stimulation with HMGB1 (500 ng/ml), the cell lysate was analyzed for the luciferase activity. Results: The expression of MIP-2, which peaked at 8h with LPS stimulation, increased sequentially until 24h after HMGB1 stimulation. An intraperitoneal injection of HMGB1, which induced a minimal increased in $IL-1{\beta}$ expression, provoked the accumulation of neutrophils the lung. A mutation of AP-1 as well as $NF-{\kappa}B$ in the IL-8 promoter region resulted in a lower luciferase activity after HMGB1 stimulation. Conclusion: The proinflammatory effects of HMGB1, particularly on IL-8, are mediated by both $NF-{\kappa}B$ and AP-1.

• Journal of Life Science
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• v.31 no.12
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• pp.1110-1119
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• 2021

The purpose of this study was to investigate whether the inflammatory response in lipopolysaccharide (LPS)-treated RAW 264.7 macrophages could be promoted by particulate matter 2.5 (PM_2.5) stimulation. To this end, the levels of inflammatory parameters, reactive oxygen species (ROS) and inflammation-regulating genes were investigated in RAW 264.7 cells treated with PM_2.5 in the presence or absence of LPS. Our results showed that the production levels of pro-inflammatory mediators (nitric oxide and prostaglandin E₂) and cytokines (interleukin-6 and -1β) were significantly increased by PM_2.5 stimulation in LPS-treated RAW 264.7 cells, which was correlated with increased expression genes involved in their production. In addition, when LPS-treated RAW 264.7 cells were exposed to PM_2.5, nuclear factor-kappaB (NF-κB) expression was further increased in the nucleus, and the expression of inhibitor of NF-κB as well as NF-κB in the cytoplasm was decreased. These results suggest that the co-treatment of PM_2.5 and LPS further increases the activation of the NF-κB signaling pathway compared to each treatment alone, thereby contributing to the promotion of transcriptional activity of inflammatory genes. Furthermore, although the generation of ROS was greatly increased by PM_2.5 in LPS-treated RAW 264.7 cells, the NF-κB inhibitor did not reduce the generation of ROS. In addition, when the generation of ROS was artificially suppressed, the production of inflammatory mediators and the activation of NF-κB were both abolished. Therefore, our results suggest that the increase in the NF-κB-mediated inflammatory response induced by PM_2.5 in LPS-treated RAW 264.7 macrophages was a ROS generation-dependent phenomenon.

• Tuberculosis and Respiratory Diseases
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• v.53 no.5
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• pp.519-529
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• 2002

Background : The therapeutic effects of surfactants on acute lung injury derive not only from their recruiting action on collapsed alveoli but also from their anti-inflammatory action in the alveolar sapce. This study evaluated the anti-inflammatory action of a surfactant in an acute lung injury model of rats by neutrophils were recollected from the BAL fluid and the NF-${\kappa}B$ activity of the neutrophilic nuclear protein was evaluated. Methods : Male Sprague-Dawley rats weighing approximately 300 gram were divided into 3 groups, which consisted of 6 rats respectively. In the control group, normal saline(3ml/kg) was instilled into the trachea twice with 30 minute interval. In two other groups, acute lung injury was induced by the intra-tracheal instillation of LPS(5mg/kg). Thirty minutes later, either a surfactant(ST group; 30mg/kg) or normal saline(NT group: 3ml/kg) was instilled via the trachea. Twenty-four hours after the LPS instillation, the BAL fluid was retrieved to measure the WBC count and cytokine(IL-$1{\beta}$ and IL-6) levels. The neutrophils were isolated from the BAL fluid and the nuclear protein was extracted to evaluate the NF-${\kappa}B$ activity using a eletrophoretic mobility shift assay(EMSA). Results : The WBC count of the BAL fluid of the ST group($3,221{\pm}1,914{\times}10^3/{\mu}l$) was higher than that of the control group($356{\pm}275{\times}10^3/{\mu}l$)(p<0.05) and lower than that of the NT group($5,561{\pm}1,757{\times}10^3/{\mu}l$)(p<0.05)). The BAL fluid level of IL-$1{\beta}$ from the NT group($2,064{\pm}1,082pg/ml$) was higher than those of the ST group($360{\pm}234pg/ml$)(p<0.05) and the control group(0pg/ml)p<0.05) and control group($49{\pm}62pg/ml$)(p<0.05). The NF-${\kappa}B$ activity of the neutrophilic nuclear protein in the ST group and NT group was similar. Conclusion : The surfactant, attenuates the alveolar inflammation in the acute lung injury of rats model. However, its anti-inflammatory action does no't appear to be mediated by the inhibition of NF-${\kappa}B$ activity.

• Journal of Mushroom
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• v.19 no.3
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• pp.225-233
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• 2021

In this study, we evaluated the anti-inflammatory effect of extract from Cudrania tricuspidata twig sawdust fermented with Ganoderma lucidum mycelium. Fermented Cudrania tricuspidata twig sawdust extracted with 70% ethanol and elucidated the potential signaling pathway in lipopolysaccharide (LPS)-induced RAW264.7 cells. Fermented Cudrania tricuspidata twig sawdust inhibits LPS-stimulated nitric oxide (NO) production without affecting cell viability in a dose-dependent manner and production of LPS-induced pro-inflammatory cytokines such as interleukin (IL)-1β, tumor necrosis factor (TNF)-α and prostaglandin2 (PGE₂). Fermented Cudrania tricuspidata twig sawdust also suppressed the expression of the pro-inflammatory mediators such as inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in LPS-stimulated RAW264.7 cells. Moreover, Fermented Cudrania tricuspidata twig sawdust significantly attenuated LPS-induced IkappaB (IκB) degradation and suppressed nuclear factor kappa B (NF-κB) nuclear translocation. These results suggest that fermented Cudrania tricuspidata twig sawdust may have great potential for the development of anti-inflammatory agent.

• Journal of the Korean Chemical Society
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• v.51 no.3
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• pp.251-257
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• 2007

The Tumor necrosis factor-α, (TNF-α), is involved in the pathogenesis of multiple sclerosis and contributes to the degeneration of oligodendrocytes as well as neurons. Nicotine has been found to have immunosuppressive and inflammation-suppressing effects. Astrocytes, the major glial cells in the CNS, are capable of producing TNF-α at both the mRNA and protein levels in response to interleukin-1 (IL-1) or TNF-α. Nicotine has been shown to influence glial cell functions. To order to explore the role of astrocytes in the production of TNF-α, astrocytes were pretreated with nicotine and are stimulated with IL-1β to determine their effects on TNF-α production. The results are as follows. Cytotoxic effects of nicotine on human fetal astrocytes were noted above 10 μg/ml of nicotine. The effect of IL-1β on TNF-α mRNA expression in primary cultured human fetal astrocytes was maximal at 2 h after IL- 1β(100 pg/ml) treatment. Human fetal astrocytes were pretreated with 0.1, 1, and 10 μg/ml of nicotine and then stimulated with IL-1β (100 pg/ml) for 2 h. The inhibitory effect of nicotine on expressions of TNF-α mRNA in human fetal astrocytes with pretreated 0.1 μg/ml of nicotine is first noted at 8 hr, and the inhibitory effect is maximal at 12 h. The inhibitory effect at 1 μg/ml of nicotine is inhibited maximal at 24 h. Nicotine at 0.1, 1 and 10 μg/ml concentrations significantly inhibits IL-1β-induced NF-κB activation. Collectively, this study indicates that nicotine might inhibit the expression of TNF-α in activated human fetal astrocytes.

• Korean Journal of Poultry Science
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• v.46 no.1
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• pp.31-37
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• 2019

A chicken clathrin-associated adaptor protein $3-{\delta}$ subunit 2 (AP3S2) is a subunit of AP3, which is involved in cargo protein trafficking to target membrane with clathrin-coated vesicles. AP3S2 may play a role in virus entry into host cells through clathrin-dependent endocytosis. AP3S2 is also known to participate in metabolic disease developments of progressions, such as liver fibrosis with hepatitis C virus infection and type 2 diabetes mellitus. Chicken AP3S2 (chAP3S2) gene was originally identified as one of the differentially expressed genes (DEGs) in chicken kidney which was fed with different calcium doses. This study aims to characterize the molecular characteristics, gene expression patterns, and transcriptional regulation of chAP3S2 in response to the stimulation of Toll-like receptor 3 (TLR3) to understand the involvement of chAP3S2 in metabolic disease in chicken. As a result, the structure prediction of chAP3S2 gene revealed that the gene is highly conserved among AP3S2 orthologs from other species. Evolutionarily, it was suggested that chAP3S2 is relatively closely related to zebrafish, and fairly far from mammal AP3S2. The transcriptional profile revealed that chAP3S2 gene was highly expressed in chicken lung and spleen tissues, and under the stimulation of poly (I:C), the chAP3S2 expression was down-regulated in DF-1 cells (P<0.05). However, the presence of the transcriptional inhibitors, BAY 11-7085 (Bay) as an inhibitor for nuclear factor ${\kappa}B$ ($NF{\kappa}B$) or Tanshinone IIA (Tan-II) as an inhibitor for activated protein 1 (AP-1), did not affect the expressional level of chAP3S2, suggesting that these transcription factors might be dispensable for TLR3 mediated repression. These results suggest that chAP3S2 gene may play a significant role against viral infection and be involved in TLR3 signaling pathway. Further study about the transcriptional regulation of chAP3S2 in TLR3 pathways and the mechanism of chAP3S2 upon virus entry shall be needed.

• Journal of the Korean Society of Food Science and Nutrition
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• v.40 no.6
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• pp.767-774
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• 2011

Defatted green tea seed was extracted with 100% ethanol for 4 hr and then fractionated with petroleum ether, ethyl acetate and butanol. The ethanol and butanol extracts showed greater increases in antiproliferation potential against liver cancer cells than petroleum ether, ethyl acetate, $H_2O$, and hot water extracts did. Thus, this study was carried out to investigate the anti-proliferative actions of defatted green tea seed ethanol extract (DGTSE) in HepG2 cancer cells. The DGTSE contained catechins including EGC ($1039.1{\pm}15.2\;g/g$), tannic acid ($683.5{\pm}17.61\;{\mu}g/g$), EC ($62.4{\pm}5.00\;{\mu}g/g$), ECG ($24.4{\pm}7.81\;{\mu}g/g$), EGCG ($20.9{\pm}0.96\;{\mu}g/g$) and gallic acid ($2.4{\pm}0.68\;{\mu}g/g$), but caffeic acid was not detected when analyzed by HPLC. The anti-proliferation effect of DGTSE toward HepG2 cells was 83.13% when treated at $10\;{\mu}g$/mL, of DGTSE, offering an $IC_{50}$ of $6.58\;{\mu}g$/mL. DGTSE decreased CYP1A1 and CYP1A2 protein expressions in a dose-dependent manner. Quinone reductase and antioxidant response element (ARE)-luciferase activities were increased about 2.6 and 1.94-fold at a concentration of $20\;{\mu}g$/mL compared to a control group, respectively. Enhancement of phase II enzyme activity by DGTSE was shown to be mediated via interaction with ARE sequences in genes encoding the phase enzymes. DGTSE significantly (p<0.05) suppressed prostaglandin $E_2$ level, tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$) protein expressions, and NF${\kappa}$B translocation, but did not affected nitric oxide production. From the above results, it is concluded that DGTSE may ameliorate tumor and inflammatory reactions through the elevation of phase II enzyme activities and suppression of NF${\kappa}$B translocation and TNF-${\alpha}$ protein expressions, which support the cancer cell anti-proliferative effects of DGTSE in HepG2 cells.

• Korean Journal of Clinical Laboratory Science
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• v.50 no.3
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• pp.337-344
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• 2018

Hepatocellular carcinoma (HCC), a major type of hepatoma, is associated with high recurrence and mortality because of its uncontrolled metastatic feature. Silymarin is a polyphenolic flavonoid from Silybum marianun (milk thistle) and exhibits anti-carcinogenic activity through modulation of the epithelial-mesenchymal transition (EMT) in several cancer cells. In this study, the inhibitory mechanism of silymarin against migration and invasion was investigated in the Huh7 HCC cell line. Wound healing and in vitro invasion assays were conducted to examine the effects of silymarin on migration and invasion. Western blot analysis was also applied to evaluate the inhibitory effects of silymarin on the EMT-related genes and their upstream signaling molecules. Silymarin inhibited the migratory and invasive activities of Huh7 cells. In addition, silymarin attenuated the protein expression levels of vimentin and matrix metalloproteinase (MMP)-9 as well as their transcription factors, Snail, and nuclear factor $(NF)-{\kappa}B$, while the expression of E-cadherin was increased by the silymarin treatment. Among the upstream signaling molecules, the phosphorylation of Akt was inhibited by the silymarin treatment, which was confirmed by the selective inhibitor, LY294002. Consequently, silymarin inhibited the invasive and migratory activities in Huh7 cells through the modulation of EMT-related gene expression by the PI3K/Akt signaling pathway, which may have potential as a chemopreventive agent against HCC metastasis.

• The Korean Journal of Food And Nutrition
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• v.29 no.3
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• pp.431-437
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• 2016

Macrophages play a pivotal role in the innate and adaptive immune systems. This study investigated the immuno-modulatory activities of polysaccharides separated from Chrysanthemum zawadskii var. latilobum (CZPS) in macrophages. Polysaccharides from Chrysanthemum zawadskii var. latilobum were extracted by the ethanol precipitation method. RAW 264.7 mouse macrophage cell line was treated with CZPS (4 to $128{\mu}g/mL$), and there was no cytotoxicity at a dose below $32{\mu}g/mL$. The levels of nitric oxide (NO) and pro-inflammatory cytokines (tumor necrosis factor (TNF)-${\alpha}$ and interleukin (IL)-6, IL-$1{\beta}$) production in the CZPS treated group ($32{\mu}g/mL$) were $6.5{\pm}0.12{\mu}m$ (NO), $1252.8{\pm}79.85$ (TNF-${\alpha}$), $305.4{\pm}29.41$ (IL-6), and $683.3{\pm}59.71$ (IL-$1{\beta}$), respectively, and they were significantly increased when compared to the control group; $2.2{\pm}0.03{\mu}m$ (NO), $452.3{\pm}38.34$ (TNF-${\alpha}$), $31.7{\pm}5.75$ (IL-6), and $184.1{\pm}11.52$ (IL-$1{\beta}$). Additionally, protein expression of inducible nitric oxide synthase (iNOS) and phosphorylation of MAPKs and NF-${\kappa}B$ expression were significantly increased upon CZPS treatment. Therefore, these results indicated that polysaccharides separated from Chrysanthemum zawadskii var. latilobum (CZPS) may have a potential immunomodulatory activity in macrophages through MAPKs and NF-${\kappa}B$ signaling, and this information is useful for the development of immune enhancing adjuvant materials using a natural ingredient.

• Journal of Life Science
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• v.27 no.11
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• pp.1256-1261
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• 2017

${\beta}$-glucan is a constituent of the cell wall of fungi, yeast and plants. It plays an important role in the immune system such as activation of immunocyte, release of pro-inflammatory and anti-cancer effect. The immune system maintains a healthy immune homeostasis. However, when pathogenic substances enter the body, immune homeostasis can break down and disease can be triggered. Therefore, we studied a substance that regulates immune homeostasis. The purpose of the study we demonstrated whether the ${\beta}$-glucan can be applied to the immune-modulation effects in human monocytic THP-1 cells. ${\beta}$-glucan was incubated in THP-1 cells at various concentrations. The $TNF-{\alpha}$ mRNA expression and protein levels were analyzed by ELISA and Real-time PCR. Additionally, the expression of MAPKs (p38 and JNK), $I{\kappa}B-{\alpha}$ and $NF-{\kappa}B$ p50 were analyzed by western blot. ${\beta}$-glucan enhanced the production of $TNF-{\alpha}$ mRNA expression and protein levels in human monocytic THP-1 cells. In addition, activation of MAPKs (p38 and JNK) and $NF-{\kappa}B$ p50 induced by ${\beta}$-glucan were increased. The study suggests that ${\beta}$-glucan contributes to immune-stimulation effect by production $TNF-{\alpha}$ in human monocytic THP-1 cells, and that MAPKs and $NF-{\kappa}B$ p50 are involved in the process. Synthetically, we have suggested ${\beta}$-glucan may be improved to immune system effect in human monocytic THP-1 cells.