• BMB Reports
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• v.50 no.2
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• pp.55-57
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• 2017

Toll-like receptor 4 (TLR4) together with MD2, one of the key pattern recognition receptors for a pathogen-associated molecular pattern, activates innate immunity by recognizing lipopolysaccharide (LPS) of Gram-negative bacteria. Although LBP and CD14 catalyze LPS transfer to the TLR4/MD2 complex, the detail mechanisms underlying this dynamic LPS transfer remain elusive. Using negative-stain electron microscopy, we visualized the dynamic intermediate complexes during LPS transfer-LBP/LPS micelles and ternary CD14/LBP/LPS micelle complexes. We also reconstituted the entire cascade of LPS transfer to TLR4/MD2 in a total internal reflection fluorescence (TIRF) microscope for a single molecule fluorescence analysis. These analyses reveal longitudinal LBP binding to the surface of LPS micelles and multi-round binding/unbinding of CD14 to single LBP/LPS micelles via key charged residues on LBP and CD14. Finally, we reveal that a single LPS molecule bound to CD14 is transferred to TLR4/MD2 in a TLR4-dependent manner. These discoveries, which clarify the molecular mechanism of dynamic LPS transfer to TLR4/MD2 via LBP and CD14, provide novel insights into the initiation of innate immune responses.

• Asian Pacific Journal of Cancer Prevention
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• v.14 no.4
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• pp.2295-2299
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• 2013

Background: To explore the role of caveolin-1(CAV-1) gene silencing on MAPK activation in lipopolysaccharide (LPS)-challenged human mammary epithelial cells. Methods: We established a MCF-10ACE of CAV-1 gene silencing from human mammary epithelial cell line MCF-10A by RNAi technology. DNA Microarray were used to detect the expression of inflammation-associated genes in MCF10ACE. Western blotting was used to examine the activation of MAPK in lipopolysaccharide(LPS)-challenged MCF-10A and MCF-10ACE. Moreover, immunofluorescence and Western bloting were performed to detect the co-localization of CAV-1 and toll-like receptor 4 (TLR4) in human mammary epithelial cells. Results: MCF-10ACE exhibited significant increases in inflammation-associated gene expression, especially IL-6 (~7-fold) and IL6R (~17-fold). In addition, LPS-induced p38 MAPK and JNK MAPK activation was significantly increased in MCF-10ACE. Furthermore, CAV-1 co-localized with TLR4 and appeared a negative correlation trend. Conclusion: CAV-1 gene silencing promotes MAPK activation via TLR4 signaling in human mammary epithelial cells response to LPS.

• Asian-Australasian Journal of Animal Sciences
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• v.31 no.1
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• pp.138-148
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• 2018

Objective: Fusarium mycotoxins deoxynivalenol (DON) and zearalenone (ZEN), common contaminants in the feed of farm animals, cause immune function impairment and organ inflammation. Consequently, the main objective of this study was to elucidate DON and ZEN effects on the mRNA expression of pro-inflammatory cytokines and other immune related genes in the kidneys of piglets. Methods: Fifteen 6-week-old piglets were randomly assigned to three dietary treatments for 4 weeks: control diet, and diets contaminated with either 8 mg DON/kg feed or 0.8 mg ZEN/kg feed. Kidney samples were collected after treatment, and RNA-seq was used to investigate the effects on immune-related genes and gene networks. Results: A total of 186 differentially expressed genes (DEGs) were screened (120 upregulated and 66 downregulated). Gene ontology analysis revealed that the immune response, and cellular and metabolic processes were significantly controlled by these DEGs. The inflammatory stimulation might be an effect of the following enriched Kyoto encyclopedia of genes and genomes pathway analysis found related to immune and disease responses: cytokine-cytokine receptor interaction, chemokine signaling pathway, toll-like receptor signaling pathway, systemic lupus erythematosus (SLE), tuberculosis, Epstein-Barr virus infection, and chemical carcinogenesis. The effects of DON and ZEN on genome-wide expression were assessed, and it was found that the DEGs associated with inflammatory cytokines (interleukin 10 receptor, beta, chemokine [C-X-C motif] ligand 9, CXCL10, chemokine [C-C motif] ligand 4), proliferation (insulin like growth factor binding protein 4, IgG heavy chain, receptor-type tyrosine-protein phosphatase C, cytochrome P450 1A1, ATP-binding cassette sub-family 8), and other immune response networks (lysozyme, complement component 4 binding protein alpha, oligoadenylate synthetase 2, signaling lymphocytic activation molecule-9, ${\alpha}$-aminoadipic semialdehyde dehydrogenase, Ig lambda chain c region, pyruvate dehydrogenase kinase, isozyme 4, carboxylesterase 1), were suppressed by DON and ZEN. Conclusion: In summary, our results indicate that high concentrations of DON and ZEN suppress the inflammatory response in kidneys, leading to potential effects on immune homeostasis.

• Molecular & Cellular Toxicology
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• v.4 no.3
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• pp.183-191
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• 2008

Volatile organic compounds (VOCs) are common constituents of cleaning and degreasing agents, paints, pesticides, personal care products, gasoline and solvents. And VOCs are evaporated at room temperature and most of them exhibit acute and chronic toxicity to human. Benzene is the most widely used prototypical VOC and the toxic mechanisms of them are still unclear. The multi-step process of toxic mechanism can be more fully understood by characterizing gene expression changes induced in cells by toxicants. In this study, DNA microarray was used to monitor the expression levels of genes in HepG2 cells and HL-60 cells exposed to the benzene on IC20 and IC50 dose respectively. In the clustering analysis of gene expression profiles, although clusters of HepG2 and HL-60 cells by benzene were divided differently, expression pattern of many genes observed similarly. We identified 916 up-regulated genes and 1,144 down-regulated genes in HepG2 cells and also 1,002 up-regulated genes and 919 down-regulated genes in HL-60 cells. The gene ontology analysis on genes expressed by benzene in HepG2 and HL-60 cells, respectively, was performed. Thus, we found some principal pathways, such as, focal adhesion, gap junction and signaling pathway in HepG2 cells and toll-like receptor signaling pathway, MAPK signaling pathway, p53 signaling pathway and neuroactive ligand-receptor interaction in HL-60 cells. And we also found 16 up-regulated and 14 down-regulated commonly expressed total 30 genes that belong in the same biological process like inflammatory response, cell cycle arrest, cell migration, transmission of nerve impulse and cell motility in two cell lines. In conclusion, we suggest that this study is meaningful because these genes regarded as strong potential biomarkers of benzene independent of cell type.

• Journal of Microbiology and Biotechnology
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• v.23 no.7
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• pp.1023-1030
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• 2013

Lipoteichoic acid (LTA), uniquely expressed on gram-positive bacteria, is recognized by Toll-like receptor 2 (TLR2) on not only antigen-presenting cells but also activated T cells. Therefore, it is reasonable to assume that LTA is acting on T cells. However, little is known about the effect of LTA on T-cell regulation. In the present study, we investigated the immunomodulatory effects of LTA on $CD4^+$ T cells. Effector $CD4^+$ T cells, induced after co-culture with S. aureus-pulsed dendritic cells, produced high levels of interferon-${\gamma}$, CD25, CD69, and TLRs 2 and 4. When effector $CD4^+$ T cells were treated with LTA, the expressions of the membrane-bound form of transforming growth factor (TGF)-${\beta}$ and forkhead box P3 increased. Coincidently, the proliferation of effector $CD4^+$ T cells was declined after LTA treatment. When TGF-${\beta}$ signaling was blocked by the TGF-${\beta}$ receptor 1 kinase inhibitor, LTA failed to suppress the proliferation of effector $CD4^+$ T cells. Therefore, the present results suggest that LTA suppresses the activity of effector $CD4^+$ T cells by enhancing TGF-${\beta}$ production.

• Asian Pacific Journal of Cancer Prevention
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• v.15 no.5
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• pp.2329-2334
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• 2014

Curcumin can inhibit proliferation of liver cancer cells by inducing apoptosis, but the specific signaling pathways involved are not completely clear. Here, we report that curcumin inhibited proliferation of MHCC97H liver cancer cells by induction of apoptosis in a concentration dependent manner via stimulating intracellular reactive oxygen species (ROS) generation. Also, we showed that increased intracellular ROS formation activated the TLR-4/MyD-88 signaling pathway, resulting in activation of caspase-8 and caspase-3, which eventually led to apoptosis in MHCC97H cells. These results showed that as an prooxidant, curcumin exerts anti-cancer effects by inducing apoptosis via the TLR-4/MyD-88 signaling pathway.

• The Journal of Internal Korean Medicine
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• v.28 no.3
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• pp.442-452
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• 2007

Objectives : The purpose of this study was to investigate the toll-like receptor (TLR)-4 mediated anti-inflammatory effects of extract from Shigyungbanha-tang (SBT) on the peritoneal macrophage. Methods : To evaluate of TLR-4 mediated inflammatory of SBT. we examined NO and cytokine production in TRL-4 ligand (LPS : lipopolysaccharide) induced macrophages. Furthermore, we examined its molecular mechanism using western blot. Results : Extract from SBT itself does not have any cytotoxic effect in the peritoneal macrophages. Extract from SBT reduced LPS-induced nitric oxide (NO). tumor necrosis factor-alpha ($TNF-{\alpha}$), interleukin (IL)-6 and IL-12 production in peritoneal macrophages. SBT inhibited degradation of inhibitor kappa B-alpha ($I{\kappa}B-{\alpha}$) in the TLR-4 mediated peritoneal macrophages. Conclusions : These results suggest that SBT inhibits NO and cytokines production through inhibiting nuclear factor-kappaB (NF-${\kappa}$B) activation in peritoneal macrophage and that SBT may be beneficial oriental medicine for inflammation.

• Journal of Physiology & Pathology in Korean Medicine
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• v.36 no.1
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• pp.28-34
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• 2022

Echinacea purpurea (Asteraceae family) is widely used in the European countries and the United States due to its proven immune enhancement and anti-inflammatory effects. Echinacea purpurea has been reported prevent and treat upper respiratory tract infections and common cold, but the underlying molecular mechanisms are not well understood. In the present study, we examined the anti-inflammatory effects and molecular mechanisms of Echinacea purpurea (EP) extract using lipopolysaccharide (LPS)-stimulated signal pathways in RAW264.7 cells. Our results suggest that EP extract exerts anti-inflammatory effects by down-regulating the expression of LPS-induced toll-like receptor 4 (TLR4), subsequently inhibiting the activation of nuclear factor (NF)-κB and mitogen-activated protein kinase (MAPK) signaling pathways and suppression of the release of pro-inflammatory cytokines. These results suggest that EP extract is a potential therapeutic agent for inflammatory diseases.

• The Korean Journal of Food And Nutrition
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• v.24 no.2
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• pp.268-272
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• 2011

전곡류의 섭취와 만성질환의 유병율은 음의 상관관계가 있는 것으로 알려져 있다. 본 연구에서는 선행 연구 결과, 지방축적억제능이 우수한 소재로 선정된 기장의 항염증능 여부를 검증하고자 하였다. 이를 위해 RAW264.7 세포에 기장열수분획($1\;{\mu}g/m\ell$ 및 $10\;{\mu}g/m\ell$)과 lipopolysaccharide(LPS)를 함께 처리한 후, 24시간 배양시켜 염증매개인자들의 분비량 및 mRNA 발현 정도를 측정하였다. 또한 LPS 자극에 대한 첫 번째 신호전달인자로 알려져 있는 interleukin-1 receptor associated kinase-4(IRAK-4)의 단백질 발현 정도를 측정하였다. 본 연구결과, 기장의 열수분획($10\;{\mu}g/m\ell$)은 LPS로 유도된 NO, $PGE_2$, TNF-${\alpha}$, IL-6 및 MCP-1의 생성량 및 mRNA 발현량을 유의적으로 억제하였다(p<0.05). 특히 이들 지표 중 pro-inflammatory cytokine인 TNF-${\alpha}$와 IL-6의 mRNA 발현량이 효과적으로 감소하였다(p<0.01). IRAK-4의 단백질 발현량 또한 유의적으로 감소하여 LPS 자극에 대한 기장열수분획의 항염증능은 toll-like receptor(TLR)를 통한 IRAK-4를 매개로 하는 신호전달체계 조절에 기인하는 것으로 사료된다.

• The Journal of Pediatrics of Korean Medicine
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• v.37 no.4
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• pp.25-33
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• 2023

Objective This study aimed to confirm the effect of Sabaek-san extract on the recovery of skin damage in atopic dermatitis-induced mice. Methods In this study, we used 4-week-old NC/Nga mice that were assigned to four groups: control (Ctrl), lipid barrier elimination (LBEG), dexamethasone (Dx) administration after lipid barrier elimination (DxAG), and Sabaek-san extract administration after lipid barrier elimination (SBAG). Ten rats were assigned to each treatment group. After drug administration for 3 days following lipid barrier elimination, ceramide kinase, caspase 14, sodium hydrogen antiporter (NHE), cathelicidin, claudin, and Toll-like receptor (TLR2) were observed to confirm restoration of skin moisturizer production, antimicrobial barriers, and tight junctions in the skin barrier. Results Ceramide kinase and caspase 14 positive reactions were significantly higher in the SBAG group than in the LBEG or DxAG groups. NHE and cathelicidin showed a higher positive reaction in the SBAG group than in the LBEG and DxAG groups. Claudins and TLR2 showed a higher positive reaction in the SBAG group than in the LBEG or DxAG groups. Conclusion It was confirmed that Sabaek-san extract may have the potential to restore damaged skin barrier in atopic dermatitis.