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Lymphotoxin β receptor signaling induces IL-8 production in human bronchial epithelial cells.

Mikami Y, Matsuzaki H, Horie M, Noguchi S, Jo T, Narumoto O, Kohyama T, Takizawa H, Nagase T, Yamauchi Y - PLoS ONE (2014)

Bottom Line: LIGHT also induced luciferase activity of NF-κB response element, but not of activator protein-1 or serum response element.Specific inhibitors of phosphorylation of extracellular signal-regulated kinase (Erk) and that of inhibitor κB attenuated IL-8 production, suggesting that LIGHT-LTβR signaling induces IL-8 production via the Erk and NF-κB pathways.LIGHT, via LTβR signaling, may contribute to exacerbation of airway neutrophilic inflammation through cytokine and chemokine production by bronchial epithelial cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Respiratory Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.

ABSTRACT
Asthma-related mortality has been decreasing due to inhaled corticosteroid use, but severe asthma remains a major clinical problem. One characteristic of severe asthma is resistance to steroid therapy, which is related to neutrophilic inflammation. Recently, the tumor necrosis factor superfamily member (TNFSF) 14/LIGHT has been recognized as a key mediator in severe asthmatic airway inflammation. However, the profiles and intracellular mechanisms of cytokine/chemokine production induced in cells by LIGHT are poorly understood. We aimed to elucidate the molecular mechanism of LIGHT-induced cytokine/chemokine production by bronchial epithelial cells. Human bronchial epithelial cells express lymphotoxin β receptor (LTβR), but not herpesvirus entry mediator, which are receptors for LIGHT. LIGHT induced various cytokines/chemokines, such as interleukin (IL)-6, oncostatin M, monocyte chemotactic protein-1, growth-regulated protein α and IL-8. Specific siRNA for LTβR attenuated IL-6 and IL-8 production by BEAS-2B and normal human bronchial epithelial cells. LIGHT activated intracellular signaling, such as mitogen-activated protein kinase and nuclear factor-κB (NF-κB) signaling. LIGHT also induced luciferase activity of NF-κB response element, but not of activator protein-1 or serum response element. Specific inhibitors of phosphorylation of extracellular signal-regulated kinase (Erk) and that of inhibitor κB attenuated IL-8 production, suggesting that LIGHT-LTβR signaling induces IL-8 production via the Erk and NF-κB pathways. LIGHT, via LTβR signaling, may contribute to exacerbation of airway neutrophilic inflammation through cytokine and chemokine production by bronchial epithelial cells.

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Time-course and dose-dependent effects of LIGHT on bronchial epithelial (BEAS-2B) cells.(A) BEAS-2B cells were stimulated with 50 ng/ml LIGHT and examined for the time-course effect on expression of mRNA for each of IL-8, IL-6, MCP-1 and RANTES. LIGHT significantly induced mRNA for each of IL-6, IL-8 and MCP-1. n = 4 separate experiments. *: p<0.05, **: p<0.01 vs 0 h. (B) BEAS-2B cells were stimulated with various concentrations of LIGHT (0, 1, 10, 100 ng/ml) for 1 h and evaluated for expression of mRNA for each of IL-8 and IL-6. LIGHT induced IL-8 and IL-6 mRNA dose-dependently. n = 4 separate experiments. ** and ††: p<0.01 vs 0 ng/ml. (C) BEAS-2B cells were stimulated with 100 ng/ml LIGHT for 24 h, and the protein concentrations in the cell supernatants were determined by ELISA. LIGHT significantly induced IL-8 and IL-6 proteins. n = 6 separate experiments. **: p<0.01.
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pone-0114791-g004: Time-course and dose-dependent effects of LIGHT on bronchial epithelial (BEAS-2B) cells.(A) BEAS-2B cells were stimulated with 50 ng/ml LIGHT and examined for the time-course effect on expression of mRNA for each of IL-8, IL-6, MCP-1 and RANTES. LIGHT significantly induced mRNA for each of IL-6, IL-8 and MCP-1. n = 4 separate experiments. *: p<0.05, **: p<0.01 vs 0 h. (B) BEAS-2B cells were stimulated with various concentrations of LIGHT (0, 1, 10, 100 ng/ml) for 1 h and evaluated for expression of mRNA for each of IL-8 and IL-6. LIGHT induced IL-8 and IL-6 mRNA dose-dependently. n = 4 separate experiments. ** and ††: p<0.01 vs 0 ng/ml. (C) BEAS-2B cells were stimulated with 100 ng/ml LIGHT for 24 h, and the protein concentrations in the cell supernatants were determined by ELISA. LIGHT significantly induced IL-8 and IL-6 proteins. n = 6 separate experiments. **: p<0.01.

Mentions: To determine the effect of LIGHT on BEAS-2B cells, we analyzed the time-course effect on expression of mRNA for several inflammatory chemokines. As shown in Fig. 4A, LIGHT significantly induced IL-8, IL-6 and MCP-1 mRNA at 1 h after stimulation. Then the cells were stimulated with several concentrations of LIGHT, and IL-8 and IL-6 mRNA expression was measured after 1 h. LIGHT induced IL-8 and IL-6 mRNA dose-dependently (Fig. 4B). In addition, as shown in Fig. 4C, LIGHT had significantly induced IL-8 and IL-6 proteins in BEAS-2B cells at 24 h after stimulation. We also confirmed by MTT assay that LIGHT, at less than 100 ng/ml, did not show any cytotoxic effect on BEAS-2B cells (data not shown). As shown in Fig. 4A, LIGHT induced clear IL-8 production at a concentration of 50 ng/ml. In consideration of that result, we used LIGHT at that concentration for subsequent experiments.


Lymphotoxin β receptor signaling induces IL-8 production in human bronchial epithelial cells.

Mikami Y, Matsuzaki H, Horie M, Noguchi S, Jo T, Narumoto O, Kohyama T, Takizawa H, Nagase T, Yamauchi Y - PLoS ONE (2014)

Time-course and dose-dependent effects of LIGHT on bronchial epithelial (BEAS-2B) cells.(A) BEAS-2B cells were stimulated with 50 ng/ml LIGHT and examined for the time-course effect on expression of mRNA for each of IL-8, IL-6, MCP-1 and RANTES. LIGHT significantly induced mRNA for each of IL-6, IL-8 and MCP-1. n = 4 separate experiments. *: p<0.05, **: p<0.01 vs 0 h. (B) BEAS-2B cells were stimulated with various concentrations of LIGHT (0, 1, 10, 100 ng/ml) for 1 h and evaluated for expression of mRNA for each of IL-8 and IL-6. LIGHT induced IL-8 and IL-6 mRNA dose-dependently. n = 4 separate experiments. ** and ††: p<0.01 vs 0 ng/ml. (C) BEAS-2B cells were stimulated with 100 ng/ml LIGHT for 24 h, and the protein concentrations in the cell supernatants were determined by ELISA. LIGHT significantly induced IL-8 and IL-6 proteins. n = 6 separate experiments. **: p<0.01.
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Related In: Results  -  Collection

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pone-0114791-g004: Time-course and dose-dependent effects of LIGHT on bronchial epithelial (BEAS-2B) cells.(A) BEAS-2B cells were stimulated with 50 ng/ml LIGHT and examined for the time-course effect on expression of mRNA for each of IL-8, IL-6, MCP-1 and RANTES. LIGHT significantly induced mRNA for each of IL-6, IL-8 and MCP-1. n = 4 separate experiments. *: p<0.05, **: p<0.01 vs 0 h. (B) BEAS-2B cells were stimulated with various concentrations of LIGHT (0, 1, 10, 100 ng/ml) for 1 h and evaluated for expression of mRNA for each of IL-8 and IL-6. LIGHT induced IL-8 and IL-6 mRNA dose-dependently. n = 4 separate experiments. ** and ††: p<0.01 vs 0 ng/ml. (C) BEAS-2B cells were stimulated with 100 ng/ml LIGHT for 24 h, and the protein concentrations in the cell supernatants were determined by ELISA. LIGHT significantly induced IL-8 and IL-6 proteins. n = 6 separate experiments. **: p<0.01.
Mentions: To determine the effect of LIGHT on BEAS-2B cells, we analyzed the time-course effect on expression of mRNA for several inflammatory chemokines. As shown in Fig. 4A, LIGHT significantly induced IL-8, IL-6 and MCP-1 mRNA at 1 h after stimulation. Then the cells were stimulated with several concentrations of LIGHT, and IL-8 and IL-6 mRNA expression was measured after 1 h. LIGHT induced IL-8 and IL-6 mRNA dose-dependently (Fig. 4B). In addition, as shown in Fig. 4C, LIGHT had significantly induced IL-8 and IL-6 proteins in BEAS-2B cells at 24 h after stimulation. We also confirmed by MTT assay that LIGHT, at less than 100 ng/ml, did not show any cytotoxic effect on BEAS-2B cells (data not shown). As shown in Fig. 4A, LIGHT induced clear IL-8 production at a concentration of 50 ng/ml. In consideration of that result, we used LIGHT at that concentration for subsequent experiments.

Bottom Line: LIGHT also induced luciferase activity of NF-κB response element, but not of activator protein-1 or serum response element.Specific inhibitors of phosphorylation of extracellular signal-regulated kinase (Erk) and that of inhibitor κB attenuated IL-8 production, suggesting that LIGHT-LTβR signaling induces IL-8 production via the Erk and NF-κB pathways.LIGHT, via LTβR signaling, may contribute to exacerbation of airway neutrophilic inflammation through cytokine and chemokine production by bronchial epithelial cells.

View Article: PubMed Central - PubMed

Affiliation: Department of Respiratory Medicine, Graduate School of Medicine, The University of Tokyo, Tokyo, Japan.

ABSTRACT
Asthma-related mortality has been decreasing due to inhaled corticosteroid use, but severe asthma remains a major clinical problem. One characteristic of severe asthma is resistance to steroid therapy, which is related to neutrophilic inflammation. Recently, the tumor necrosis factor superfamily member (TNFSF) 14/LIGHT has been recognized as a key mediator in severe asthmatic airway inflammation. However, the profiles and intracellular mechanisms of cytokine/chemokine production induced in cells by LIGHT are poorly understood. We aimed to elucidate the molecular mechanism of LIGHT-induced cytokine/chemokine production by bronchial epithelial cells. Human bronchial epithelial cells express lymphotoxin β receptor (LTβR), but not herpesvirus entry mediator, which are receptors for LIGHT. LIGHT induced various cytokines/chemokines, such as interleukin (IL)-6, oncostatin M, monocyte chemotactic protein-1, growth-regulated protein α and IL-8. Specific siRNA for LTβR attenuated IL-6 and IL-8 production by BEAS-2B and normal human bronchial epithelial cells. LIGHT activated intracellular signaling, such as mitogen-activated protein kinase and nuclear factor-κB (NF-κB) signaling. LIGHT also induced luciferase activity of NF-κB response element, but not of activator protein-1 or serum response element. Specific inhibitors of phosphorylation of extracellular signal-regulated kinase (Erk) and that of inhibitor κB attenuated IL-8 production, suggesting that LIGHT-LTβR signaling induces IL-8 production via the Erk and NF-κB pathways. LIGHT, via LTβR signaling, may contribute to exacerbation of airway neutrophilic inflammation through cytokine and chemokine production by bronchial epithelial cells.

Show MeSH
Related in: MedlinePlus