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    Sino Biological 4 1bb fusion protein treatment100
    Blockade of <t>4-1BB</t> signaling attenuates the secretion of pro-fibrotic mediators by MH-S cells. Transfected (Len-cont. and sh-4-1BB) MH-S cells were treated with or without NQDI 1 (10 µM), 4-1BBIg (10 µg/mL), or IgG1 (10 µg/mL) for 2 h, then exposed to crystalline silica (50 µg/cm 2 ) for 12 h. (A) Western blots analysis of ASK-1 and downstream mitogen-activated protein kinase proteins (p38 and JNK/stress activated protein kinase) and their phosphorylated forms. (B–D) Levels of phospho-ASK1, phospho-p38, and phospho-JNK were normalized to those of β-actin ( n = 3). (E,G) Western blots analysis of IκBα and phospho-IκBα. (F,H) The level of phospho-IκBα was normalized to those of β-actin ( n = 3). (I–K) The expressions of MMP12, MMP9, and monocyte chemoattractant protein-1 were detected by real-time polymerase chain reaction analysis ( n = 4). ELISA analysis of cytokines in the culture supernatants. (L) IL-1β, (M) IL-6, (N) tumor necrosis factor-α ( n = 4). Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant). The data were representative of three independent experiments.
    4 1bb Fusion Protein Treatment100, supplied by Sino Biological, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/4 1bb fusion protein treatment100/product/Sino Biological
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    4 1bb fusion protein treatment100 - by Bioz Stars, 2021-09
    86/100 stars

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    1) Product Images from "4-1BB Signaling Promotes Alveolar Macrophages-Mediated Pro-Fibrotic Responses and Crystalline Silica-Induced Pulmonary Fibrosis in Mice"

    Article Title: 4-1BB Signaling Promotes Alveolar Macrophages-Mediated Pro-Fibrotic Responses and Crystalline Silica-Induced Pulmonary Fibrosis in Mice

    Journal: Frontiers in Immunology

    doi: 10.3389/fimmu.2018.01848

    Blockade of 4-1BB signaling attenuates the secretion of pro-fibrotic mediators by MH-S cells. Transfected (Len-cont. and sh-4-1BB) MH-S cells were treated with or without NQDI 1 (10 µM), 4-1BBIg (10 µg/mL), or IgG1 (10 µg/mL) for 2 h, then exposed to crystalline silica (50 µg/cm 2 ) for 12 h. (A) Western blots analysis of ASK-1 and downstream mitogen-activated protein kinase proteins (p38 and JNK/stress activated protein kinase) and their phosphorylated forms. (B–D) Levels of phospho-ASK1, phospho-p38, and phospho-JNK were normalized to those of β-actin ( n = 3). (E,G) Western blots analysis of IκBα and phospho-IκBα. (F,H) The level of phospho-IκBα was normalized to those of β-actin ( n = 3). (I–K) The expressions of MMP12, MMP9, and monocyte chemoattractant protein-1 were detected by real-time polymerase chain reaction analysis ( n = 4). ELISA analysis of cytokines in the culture supernatants. (L) IL-1β, (M) IL-6, (N) tumor necrosis factor-α ( n = 4). Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant). The data were representative of three independent experiments.
    Figure Legend Snippet: Blockade of 4-1BB signaling attenuates the secretion of pro-fibrotic mediators by MH-S cells. Transfected (Len-cont. and sh-4-1BB) MH-S cells were treated with or without NQDI 1 (10 µM), 4-1BBIg (10 µg/mL), or IgG1 (10 µg/mL) for 2 h, then exposed to crystalline silica (50 µg/cm 2 ) for 12 h. (A) Western blots analysis of ASK-1 and downstream mitogen-activated protein kinase proteins (p38 and JNK/stress activated protein kinase) and their phosphorylated forms. (B–D) Levels of phospho-ASK1, phospho-p38, and phospho-JNK were normalized to those of β-actin ( n = 3). (E,G) Western blots analysis of IκBα and phospho-IκBα. (F,H) The level of phospho-IκBα was normalized to those of β-actin ( n = 3). (I–K) The expressions of MMP12, MMP9, and monocyte chemoattractant protein-1 were detected by real-time polymerase chain reaction analysis ( n = 4). ELISA analysis of cytokines in the culture supernatants. (L) IL-1β, (M) IL-6, (N) tumor necrosis factor-α ( n = 4). Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant). The data were representative of three independent experiments.

    Techniques Used: Transfection, Western Blot, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    A model for alveolar macrophages (AMs) expressing 4-1BB in the regulation of pulmonary fibrosis through secreting pro-fibrotic mediators. The expression of 4-1BB increases in AMs in response to crystalline silica, which leads to elevated secretion of pro-inflammatory and pro-fibrotic cytokines, chemokines, and MMPs. These pro-fibrotic mediators promote pulmonary alveoli injury, the accumulation of monocytes, lymphocytes, and fibrocytes, and collagen deposition, resulting in pulmonary fibrosis.
    Figure Legend Snippet: A model for alveolar macrophages (AMs) expressing 4-1BB in the regulation of pulmonary fibrosis through secreting pro-fibrotic mediators. The expression of 4-1BB increases in AMs in response to crystalline silica, which leads to elevated secretion of pro-inflammatory and pro-fibrotic cytokines, chemokines, and MMPs. These pro-fibrotic mediators promote pulmonary alveoli injury, the accumulation of monocytes, lymphocytes, and fibrocytes, and collagen deposition, resulting in pulmonary fibrosis.

    Techniques Used: Affinity Magnetic Separation, Expressing

    Expression of 4-1BB on CD4+ T cells. (A) Representative plots of flow cytometric analyses for 4-1BB on effector and naïve T cells. (B) The percentage of effector T cells expressing 4-1BB. (C,D) The frequency of effector and naïve T cells in CD4+ T cells from the lungs. (E) The percentage of naïve T cells expressing 4-1BB. The experiments were performed twice with similar results. Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; ns, not significant).
    Figure Legend Snippet: Expression of 4-1BB on CD4+ T cells. (A) Representative plots of flow cytometric analyses for 4-1BB on effector and naïve T cells. (B) The percentage of effector T cells expressing 4-1BB. (C,D) The frequency of effector and naïve T cells in CD4+ T cells from the lungs. (E) The percentage of naïve T cells expressing 4-1BB. The experiments were performed twice with similar results. Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; ns, not significant).

    Techniques Used: Expressing

    4-1BB expression on the mouse alveolar macrophage cell line, MH-S. MH-S cells were treated with crystalline silica (50 µg/cm 2 ) or saline for 12 h. (A,B) The percentage of MH-S cells expressing 4-1BB ( n = 4). (C) Western blot analysis of 4-1BB protein from whole cell lysates. (D) Quantification of the 4-1BB protein level relative to that of β-actin is shown ( n = 3). (E) Total RNA was isolated to analyze 4-1BB mRNA expression ( n = 4) relative to GAPDH. The data were representative of three independent experiments. Data were expressed as mean ± SEM (*** p ≤ 0.001).
    Figure Legend Snippet: 4-1BB expression on the mouse alveolar macrophage cell line, MH-S. MH-S cells were treated with crystalline silica (50 µg/cm 2 ) or saline for 12 h. (A,B) The percentage of MH-S cells expressing 4-1BB ( n = 4). (C) Western blot analysis of 4-1BB protein from whole cell lysates. (D) Quantification of the 4-1BB protein level relative to that of β-actin is shown ( n = 3). (E) Total RNA was isolated to analyze 4-1BB mRNA expression ( n = 4) relative to GAPDH. The data were representative of three independent experiments. Data were expressed as mean ± SEM (*** p ≤ 0.001).

    Techniques Used: Expressing, Western Blot, Isolation

    Activation of 4-1BB signaling promotes the secretion of pro-fibrotic mediators by MH-S cells. MH-S cells treated with or without agonist 4-1BB mAb (10 µg/mL) or IgG (10 µg/mL) for 2 h prior to exposure to crystalline silica (50 µg/cm 2 ) for 12 h. (A) Western blots analysis of ASK-1 and downstream mitogen-activated protein kinase proteins (p38 and JNK/stress activated protein kinase) and their phosphorylated forms. (B–D) The levels of phospho-ASK1, phospho-p38, and phospho-JNK were normalized to those of β-actin ( n = 3). (E) Western blots analysis of IκBα and phospho-IκBα. (F) The level of phospho-IκBα was normalized to those of β-actin ( n = 3). (G–I) Real-time polymerase chain reaction analysis of MMP12, MMP9, and monocyte chemoattractant protein-1 mRNA expression ( n = 4). (J–L) ELISA analysis was used to quantify the secretion of IL-1β, IL-6 and tumor necrosis factor-α ( n = 4). The results were representative of three independent experiments. Results were graphed as the mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001).
    Figure Legend Snippet: Activation of 4-1BB signaling promotes the secretion of pro-fibrotic mediators by MH-S cells. MH-S cells treated with or without agonist 4-1BB mAb (10 µg/mL) or IgG (10 µg/mL) for 2 h prior to exposure to crystalline silica (50 µg/cm 2 ) for 12 h. (A) Western blots analysis of ASK-1 and downstream mitogen-activated protein kinase proteins (p38 and JNK/stress activated protein kinase) and their phosphorylated forms. (B–D) The levels of phospho-ASK1, phospho-p38, and phospho-JNK were normalized to those of β-actin ( n = 3). (E) Western blots analysis of IκBα and phospho-IκBα. (F) The level of phospho-IκBα was normalized to those of β-actin ( n = 3). (G–I) Real-time polymerase chain reaction analysis of MMP12, MMP9, and monocyte chemoattractant protein-1 mRNA expression ( n = 4). (J–L) ELISA analysis was used to quantify the secretion of IL-1β, IL-6 and tumor necrosis factor-α ( n = 4). The results were representative of three independent experiments. Results were graphed as the mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001).

    Techniques Used: Activation Assay, Western Blot, Real-time Polymerase Chain Reaction, Expressing, Enzyme-linked Immunosorbent Assay

    The secretion of pro-fibrotic mediators is reduced in the lungs from crystalline silica (CS)-injured mice, upon inhibition of 4-1BB signaling. C57BL/6 mice were administered a CS suspension or saline, respectively; 4-1BBIg or isotype control (IgG1) were injected intraperitoneally (i.p.; n = 3–4). (A–D) Quantification of MMP9 and MMP12 protein levels by western blot, which were normalized to those of β-actin in lungs. Shown as bar graph. (E–H) ELISA analysis of cytokines in lung tissues. (E) IL-1β, (F) IL-6, (G) tumor necrosis factor-α, (H) monocyte chemoattractant protein-1. Experiments were performed three times. C57BL/6 mice were administered a CS suspension or saline, respectively; NQDI 1 or isotype control were injected i.p. ( n = 10). (I) Immunohistochemical staining of paraffin-embedded lung tissue sections at 7 and 56 days showed CD68, MMP9, and MMP12 expression. Nuclei were stained by hematoxylin (blue). (J–L) Identification of MMP9 and MMP12 protein levels in mouse lung tissues at 7 and 56 days by western blot. The levels of MMP9 and MMP12 were normalized to those of β-actin. (M) Representative images for the immunohistochemical staining of collagen I in paraffin-embedded lung tissue sections 56 days after CS instillation. Nuclei were stained by hematoxylin (blue). (I,M) Scale bar, 50 µm. Experiments were performed three times. Data are shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant).
    Figure Legend Snippet: The secretion of pro-fibrotic mediators is reduced in the lungs from crystalline silica (CS)-injured mice, upon inhibition of 4-1BB signaling. C57BL/6 mice were administered a CS suspension or saline, respectively; 4-1BBIg or isotype control (IgG1) were injected intraperitoneally (i.p.; n = 3–4). (A–D) Quantification of MMP9 and MMP12 protein levels by western blot, which were normalized to those of β-actin in lungs. Shown as bar graph. (E–H) ELISA analysis of cytokines in lung tissues. (E) IL-1β, (F) IL-6, (G) tumor necrosis factor-α, (H) monocyte chemoattractant protein-1. Experiments were performed three times. C57BL/6 mice were administered a CS suspension or saline, respectively; NQDI 1 or isotype control were injected i.p. ( n = 10). (I) Immunohistochemical staining of paraffin-embedded lung tissue sections at 7 and 56 days showed CD68, MMP9, and MMP12 expression. Nuclei were stained by hematoxylin (blue). (J–L) Identification of MMP9 and MMP12 protein levels in mouse lung tissues at 7 and 56 days by western blot. The levels of MMP9 and MMP12 were normalized to those of β-actin. (M) Representative images for the immunohistochemical staining of collagen I in paraffin-embedded lung tissue sections 56 days after CS instillation. Nuclei were stained by hematoxylin (blue). (I,M) Scale bar, 50 µm. Experiments were performed three times. Data are shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant).

    Techniques Used: Mouse Assay, Inhibition, Injection, Western Blot, Enzyme-linked Immunosorbent Assay, Immunohistochemistry, Staining, Expressing

    Expression of 4-1BB (CD137) and 4-1BBL (CD137L) on pulmonary macrophages. After 7-days exposure to crystalline silica, mice were sacrificed. The lungs were prepared as single-cell suspensions for flow cytometric analyses ( n = 3–4). (A) Representative plots of flow cytometric analyses for 4-1BB and 4-1BBL on alveolar macrophages (AMs) and interstitial macrophages (IMs). (B,C) The percentage of AMs expressing 4-1BB and 4-1BBL. (D,E) The frequency of AMs and IMs in CD45+ cells from the lungs. (F,G) The percentage of IMs expressing 4-1BB and 4-1BBL. The experiments were performed twice with similar results. Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant).
    Figure Legend Snippet: Expression of 4-1BB (CD137) and 4-1BBL (CD137L) on pulmonary macrophages. After 7-days exposure to crystalline silica, mice were sacrificed. The lungs were prepared as single-cell suspensions for flow cytometric analyses ( n = 3–4). (A) Representative plots of flow cytometric analyses for 4-1BB and 4-1BBL on alveolar macrophages (AMs) and interstitial macrophages (IMs). (B,C) The percentage of AMs expressing 4-1BB and 4-1BBL. (D,E) The frequency of AMs and IMs in CD45+ cells from the lungs. (F,G) The percentage of IMs expressing 4-1BB and 4-1BBL. The experiments were performed twice with similar results. Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant).

    Techniques Used: Expressing, Mouse Assay, Affinity Magnetic Separation

    Related Articles

    Injection:

    Article Title: 4-1BB Signaling Promotes Alveolar Macrophages-Mediated Pro-Fibrotic Responses and Crystalline Silica-Induced Pulmonary Fibrosis in Mice
    Article Snippet: .. For 4-1BB Fusion Protein Treatment100 or 50 µg of 4-1BB fusion protein (4-1BBIg; cat: 50811-M02H, Sino Biological Inc., Beijing, China) per mouse or 100 µg isotype IgG (human IgG1; Sino Biological Inc.) were injected intraperitoneally into mice, on days 1 and 4 after CS administration (3–4 mice per group). ..

    Mouse Assay:

    Article Title: 4-1BB Signaling Promotes Alveolar Macrophages-Mediated Pro-Fibrotic Responses and Crystalline Silica-Induced Pulmonary Fibrosis in Mice
    Article Snippet: .. For 4-1BB Fusion Protein Treatment100 or 50 µg of 4-1BB fusion protein (4-1BBIg; cat: 50811-M02H, Sino Biological Inc., Beijing, China) per mouse or 100 µg isotype IgG (human IgG1; Sino Biological Inc.) were injected intraperitoneally into mice, on days 1 and 4 after CS administration (3–4 mice per group). ..

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    Sino Biological 4 1bb fusion protein treatment100
    Blockade of <t>4-1BB</t> signaling attenuates the secretion of pro-fibrotic mediators by MH-S cells. Transfected (Len-cont. and sh-4-1BB) MH-S cells were treated with or without NQDI 1 (10 µM), 4-1BBIg (10 µg/mL), or IgG1 (10 µg/mL) for 2 h, then exposed to crystalline silica (50 µg/cm 2 ) for 12 h. (A) Western blots analysis of ASK-1 and downstream mitogen-activated protein kinase proteins (p38 and JNK/stress activated protein kinase) and their phosphorylated forms. (B–D) Levels of phospho-ASK1, phospho-p38, and phospho-JNK were normalized to those of β-actin ( n = 3). (E,G) Western blots analysis of IκBα and phospho-IκBα. (F,H) The level of phospho-IκBα was normalized to those of β-actin ( n = 3). (I–K) The expressions of MMP12, MMP9, and monocyte chemoattractant protein-1 were detected by real-time polymerase chain reaction analysis ( n = 4). ELISA analysis of cytokines in the culture supernatants. (L) IL-1β, (M) IL-6, (N) tumor necrosis factor-α ( n = 4). Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant). The data were representative of three independent experiments.
    4 1bb Fusion Protein Treatment100, supplied by Sino Biological, used in various techniques. Bioz Stars score: 86/100, based on 1 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/4 1bb fusion protein treatment100/product/Sino Biological
    Average 86 stars, based on 1 article reviews
    Price from $9.99 to $1999.99
    4 1bb fusion protein treatment100 - by Bioz Stars, 2021-09
    86/100 stars
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    Blockade of 4-1BB signaling attenuates the secretion of pro-fibrotic mediators by MH-S cells. Transfected (Len-cont. and sh-4-1BB) MH-S cells were treated with or without NQDI 1 (10 µM), 4-1BBIg (10 µg/mL), or IgG1 (10 µg/mL) for 2 h, then exposed to crystalline silica (50 µg/cm 2 ) for 12 h. (A) Western blots analysis of ASK-1 and downstream mitogen-activated protein kinase proteins (p38 and JNK/stress activated protein kinase) and their phosphorylated forms. (B–D) Levels of phospho-ASK1, phospho-p38, and phospho-JNK were normalized to those of β-actin ( n = 3). (E,G) Western blots analysis of IκBα and phospho-IκBα. (F,H) The level of phospho-IκBα was normalized to those of β-actin ( n = 3). (I–K) The expressions of MMP12, MMP9, and monocyte chemoattractant protein-1 were detected by real-time polymerase chain reaction analysis ( n = 4). ELISA analysis of cytokines in the culture supernatants. (L) IL-1β, (M) IL-6, (N) tumor necrosis factor-α ( n = 4). Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant). The data were representative of three independent experiments.

    Journal: Frontiers in Immunology

    Article Title: 4-1BB Signaling Promotes Alveolar Macrophages-Mediated Pro-Fibrotic Responses and Crystalline Silica-Induced Pulmonary Fibrosis in Mice

    doi: 10.3389/fimmu.2018.01848

    Figure Lengend Snippet: Blockade of 4-1BB signaling attenuates the secretion of pro-fibrotic mediators by MH-S cells. Transfected (Len-cont. and sh-4-1BB) MH-S cells were treated with or without NQDI 1 (10 µM), 4-1BBIg (10 µg/mL), or IgG1 (10 µg/mL) for 2 h, then exposed to crystalline silica (50 µg/cm 2 ) for 12 h. (A) Western blots analysis of ASK-1 and downstream mitogen-activated protein kinase proteins (p38 and JNK/stress activated protein kinase) and their phosphorylated forms. (B–D) Levels of phospho-ASK1, phospho-p38, and phospho-JNK were normalized to those of β-actin ( n = 3). (E,G) Western blots analysis of IκBα and phospho-IκBα. (F,H) The level of phospho-IκBα was normalized to those of β-actin ( n = 3). (I–K) The expressions of MMP12, MMP9, and monocyte chemoattractant protein-1 were detected by real-time polymerase chain reaction analysis ( n = 4). ELISA analysis of cytokines in the culture supernatants. (L) IL-1β, (M) IL-6, (N) tumor necrosis factor-α ( n = 4). Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant). The data were representative of three independent experiments.

    Article Snippet: For 4-1BB Fusion Protein Treatment100 or 50 µg of 4-1BB fusion protein (4-1BBIg; cat: 50811-M02H, Sino Biological Inc., Beijing, China) per mouse or 100 µg isotype IgG (human IgG1; Sino Biological Inc.) were injected intraperitoneally into mice, on days 1 and 4 after CS administration (3–4 mice per group).

    Techniques: Transfection, Western Blot, Real-time Polymerase Chain Reaction, Enzyme-linked Immunosorbent Assay

    A model for alveolar macrophages (AMs) expressing 4-1BB in the regulation of pulmonary fibrosis through secreting pro-fibrotic mediators. The expression of 4-1BB increases in AMs in response to crystalline silica, which leads to elevated secretion of pro-inflammatory and pro-fibrotic cytokines, chemokines, and MMPs. These pro-fibrotic mediators promote pulmonary alveoli injury, the accumulation of monocytes, lymphocytes, and fibrocytes, and collagen deposition, resulting in pulmonary fibrosis.

    Journal: Frontiers in Immunology

    Article Title: 4-1BB Signaling Promotes Alveolar Macrophages-Mediated Pro-Fibrotic Responses and Crystalline Silica-Induced Pulmonary Fibrosis in Mice

    doi: 10.3389/fimmu.2018.01848

    Figure Lengend Snippet: A model for alveolar macrophages (AMs) expressing 4-1BB in the regulation of pulmonary fibrosis through secreting pro-fibrotic mediators. The expression of 4-1BB increases in AMs in response to crystalline silica, which leads to elevated secretion of pro-inflammatory and pro-fibrotic cytokines, chemokines, and MMPs. These pro-fibrotic mediators promote pulmonary alveoli injury, the accumulation of monocytes, lymphocytes, and fibrocytes, and collagen deposition, resulting in pulmonary fibrosis.

    Article Snippet: For 4-1BB Fusion Protein Treatment100 or 50 µg of 4-1BB fusion protein (4-1BBIg; cat: 50811-M02H, Sino Biological Inc., Beijing, China) per mouse or 100 µg isotype IgG (human IgG1; Sino Biological Inc.) were injected intraperitoneally into mice, on days 1 and 4 after CS administration (3–4 mice per group).

    Techniques: Affinity Magnetic Separation, Expressing

    Expression of 4-1BB on CD4+ T cells. (A) Representative plots of flow cytometric analyses for 4-1BB on effector and naïve T cells. (B) The percentage of effector T cells expressing 4-1BB. (C,D) The frequency of effector and naïve T cells in CD4+ T cells from the lungs. (E) The percentage of naïve T cells expressing 4-1BB. The experiments were performed twice with similar results. Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; ns, not significant).

    Journal: Frontiers in Immunology

    Article Title: 4-1BB Signaling Promotes Alveolar Macrophages-Mediated Pro-Fibrotic Responses and Crystalline Silica-Induced Pulmonary Fibrosis in Mice

    doi: 10.3389/fimmu.2018.01848

    Figure Lengend Snippet: Expression of 4-1BB on CD4+ T cells. (A) Representative plots of flow cytometric analyses for 4-1BB on effector and naïve T cells. (B) The percentage of effector T cells expressing 4-1BB. (C,D) The frequency of effector and naïve T cells in CD4+ T cells from the lungs. (E) The percentage of naïve T cells expressing 4-1BB. The experiments were performed twice with similar results. Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; ns, not significant).

    Article Snippet: For 4-1BB Fusion Protein Treatment100 or 50 µg of 4-1BB fusion protein (4-1BBIg; cat: 50811-M02H, Sino Biological Inc., Beijing, China) per mouse or 100 µg isotype IgG (human IgG1; Sino Biological Inc.) were injected intraperitoneally into mice, on days 1 and 4 after CS administration (3–4 mice per group).

    Techniques: Expressing

    4-1BB expression on the mouse alveolar macrophage cell line, MH-S. MH-S cells were treated with crystalline silica (50 µg/cm 2 ) or saline for 12 h. (A,B) The percentage of MH-S cells expressing 4-1BB ( n = 4). (C) Western blot analysis of 4-1BB protein from whole cell lysates. (D) Quantification of the 4-1BB protein level relative to that of β-actin is shown ( n = 3). (E) Total RNA was isolated to analyze 4-1BB mRNA expression ( n = 4) relative to GAPDH. The data were representative of three independent experiments. Data were expressed as mean ± SEM (*** p ≤ 0.001).

    Journal: Frontiers in Immunology

    Article Title: 4-1BB Signaling Promotes Alveolar Macrophages-Mediated Pro-Fibrotic Responses and Crystalline Silica-Induced Pulmonary Fibrosis in Mice

    doi: 10.3389/fimmu.2018.01848

    Figure Lengend Snippet: 4-1BB expression on the mouse alveolar macrophage cell line, MH-S. MH-S cells were treated with crystalline silica (50 µg/cm 2 ) or saline for 12 h. (A,B) The percentage of MH-S cells expressing 4-1BB ( n = 4). (C) Western blot analysis of 4-1BB protein from whole cell lysates. (D) Quantification of the 4-1BB protein level relative to that of β-actin is shown ( n = 3). (E) Total RNA was isolated to analyze 4-1BB mRNA expression ( n = 4) relative to GAPDH. The data were representative of three independent experiments. Data were expressed as mean ± SEM (*** p ≤ 0.001).

    Article Snippet: For 4-1BB Fusion Protein Treatment100 or 50 µg of 4-1BB fusion protein (4-1BBIg; cat: 50811-M02H, Sino Biological Inc., Beijing, China) per mouse or 100 µg isotype IgG (human IgG1; Sino Biological Inc.) were injected intraperitoneally into mice, on days 1 and 4 after CS administration (3–4 mice per group).

    Techniques: Expressing, Western Blot, Isolation

    Activation of 4-1BB signaling promotes the secretion of pro-fibrotic mediators by MH-S cells. MH-S cells treated with or without agonist 4-1BB mAb (10 µg/mL) or IgG (10 µg/mL) for 2 h prior to exposure to crystalline silica (50 µg/cm 2 ) for 12 h. (A) Western blots analysis of ASK-1 and downstream mitogen-activated protein kinase proteins (p38 and JNK/stress activated protein kinase) and their phosphorylated forms. (B–D) The levels of phospho-ASK1, phospho-p38, and phospho-JNK were normalized to those of β-actin ( n = 3). (E) Western blots analysis of IκBα and phospho-IκBα. (F) The level of phospho-IκBα was normalized to those of β-actin ( n = 3). (G–I) Real-time polymerase chain reaction analysis of MMP12, MMP9, and monocyte chemoattractant protein-1 mRNA expression ( n = 4). (J–L) ELISA analysis was used to quantify the secretion of IL-1β, IL-6 and tumor necrosis factor-α ( n = 4). The results were representative of three independent experiments. Results were graphed as the mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001).

    Journal: Frontiers in Immunology

    Article Title: 4-1BB Signaling Promotes Alveolar Macrophages-Mediated Pro-Fibrotic Responses and Crystalline Silica-Induced Pulmonary Fibrosis in Mice

    doi: 10.3389/fimmu.2018.01848

    Figure Lengend Snippet: Activation of 4-1BB signaling promotes the secretion of pro-fibrotic mediators by MH-S cells. MH-S cells treated with or without agonist 4-1BB mAb (10 µg/mL) or IgG (10 µg/mL) for 2 h prior to exposure to crystalline silica (50 µg/cm 2 ) for 12 h. (A) Western blots analysis of ASK-1 and downstream mitogen-activated protein kinase proteins (p38 and JNK/stress activated protein kinase) and their phosphorylated forms. (B–D) The levels of phospho-ASK1, phospho-p38, and phospho-JNK were normalized to those of β-actin ( n = 3). (E) Western blots analysis of IκBα and phospho-IκBα. (F) The level of phospho-IκBα was normalized to those of β-actin ( n = 3). (G–I) Real-time polymerase chain reaction analysis of MMP12, MMP9, and monocyte chemoattractant protein-1 mRNA expression ( n = 4). (J–L) ELISA analysis was used to quantify the secretion of IL-1β, IL-6 and tumor necrosis factor-α ( n = 4). The results were representative of three independent experiments. Results were graphed as the mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001).

    Article Snippet: For 4-1BB Fusion Protein Treatment100 or 50 µg of 4-1BB fusion protein (4-1BBIg; cat: 50811-M02H, Sino Biological Inc., Beijing, China) per mouse or 100 µg isotype IgG (human IgG1; Sino Biological Inc.) were injected intraperitoneally into mice, on days 1 and 4 after CS administration (3–4 mice per group).

    Techniques: Activation Assay, Western Blot, Real-time Polymerase Chain Reaction, Expressing, Enzyme-linked Immunosorbent Assay

    The secretion of pro-fibrotic mediators is reduced in the lungs from crystalline silica (CS)-injured mice, upon inhibition of 4-1BB signaling. C57BL/6 mice were administered a CS suspension or saline, respectively; 4-1BBIg or isotype control (IgG1) were injected intraperitoneally (i.p.; n = 3–4). (A–D) Quantification of MMP9 and MMP12 protein levels by western blot, which were normalized to those of β-actin in lungs. Shown as bar graph. (E–H) ELISA analysis of cytokines in lung tissues. (E) IL-1β, (F) IL-6, (G) tumor necrosis factor-α, (H) monocyte chemoattractant protein-1. Experiments were performed three times. C57BL/6 mice were administered a CS suspension or saline, respectively; NQDI 1 or isotype control were injected i.p. ( n = 10). (I) Immunohistochemical staining of paraffin-embedded lung tissue sections at 7 and 56 days showed CD68, MMP9, and MMP12 expression. Nuclei were stained by hematoxylin (blue). (J–L) Identification of MMP9 and MMP12 protein levels in mouse lung tissues at 7 and 56 days by western blot. The levels of MMP9 and MMP12 were normalized to those of β-actin. (M) Representative images for the immunohistochemical staining of collagen I in paraffin-embedded lung tissue sections 56 days after CS instillation. Nuclei were stained by hematoxylin (blue). (I,M) Scale bar, 50 µm. Experiments were performed three times. Data are shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant).

    Journal: Frontiers in Immunology

    Article Title: 4-1BB Signaling Promotes Alveolar Macrophages-Mediated Pro-Fibrotic Responses and Crystalline Silica-Induced Pulmonary Fibrosis in Mice

    doi: 10.3389/fimmu.2018.01848

    Figure Lengend Snippet: The secretion of pro-fibrotic mediators is reduced in the lungs from crystalline silica (CS)-injured mice, upon inhibition of 4-1BB signaling. C57BL/6 mice were administered a CS suspension or saline, respectively; 4-1BBIg or isotype control (IgG1) were injected intraperitoneally (i.p.; n = 3–4). (A–D) Quantification of MMP9 and MMP12 protein levels by western blot, which were normalized to those of β-actin in lungs. Shown as bar graph. (E–H) ELISA analysis of cytokines in lung tissues. (E) IL-1β, (F) IL-6, (G) tumor necrosis factor-α, (H) monocyte chemoattractant protein-1. Experiments were performed three times. C57BL/6 mice were administered a CS suspension or saline, respectively; NQDI 1 or isotype control were injected i.p. ( n = 10). (I) Immunohistochemical staining of paraffin-embedded lung tissue sections at 7 and 56 days showed CD68, MMP9, and MMP12 expression. Nuclei were stained by hematoxylin (blue). (J–L) Identification of MMP9 and MMP12 protein levels in mouse lung tissues at 7 and 56 days by western blot. The levels of MMP9 and MMP12 were normalized to those of β-actin. (M) Representative images for the immunohistochemical staining of collagen I in paraffin-embedded lung tissue sections 56 days after CS instillation. Nuclei were stained by hematoxylin (blue). (I,M) Scale bar, 50 µm. Experiments were performed three times. Data are shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant).

    Article Snippet: For 4-1BB Fusion Protein Treatment100 or 50 µg of 4-1BB fusion protein (4-1BBIg; cat: 50811-M02H, Sino Biological Inc., Beijing, China) per mouse or 100 µg isotype IgG (human IgG1; Sino Biological Inc.) were injected intraperitoneally into mice, on days 1 and 4 after CS administration (3–4 mice per group).

    Techniques: Mouse Assay, Inhibition, Injection, Western Blot, Enzyme-linked Immunosorbent Assay, Immunohistochemistry, Staining, Expressing

    Expression of 4-1BB (CD137) and 4-1BBL (CD137L) on pulmonary macrophages. After 7-days exposure to crystalline silica, mice were sacrificed. The lungs were prepared as single-cell suspensions for flow cytometric analyses ( n = 3–4). (A) Representative plots of flow cytometric analyses for 4-1BB and 4-1BBL on alveolar macrophages (AMs) and interstitial macrophages (IMs). (B,C) The percentage of AMs expressing 4-1BB and 4-1BBL. (D,E) The frequency of AMs and IMs in CD45+ cells from the lungs. (F,G) The percentage of IMs expressing 4-1BB and 4-1BBL. The experiments were performed twice with similar results. Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant).

    Journal: Frontiers in Immunology

    Article Title: 4-1BB Signaling Promotes Alveolar Macrophages-Mediated Pro-Fibrotic Responses and Crystalline Silica-Induced Pulmonary Fibrosis in Mice

    doi: 10.3389/fimmu.2018.01848

    Figure Lengend Snippet: Expression of 4-1BB (CD137) and 4-1BBL (CD137L) on pulmonary macrophages. After 7-days exposure to crystalline silica, mice were sacrificed. The lungs were prepared as single-cell suspensions for flow cytometric analyses ( n = 3–4). (A) Representative plots of flow cytometric analyses for 4-1BB and 4-1BBL on alveolar macrophages (AMs) and interstitial macrophages (IMs). (B,C) The percentage of AMs expressing 4-1BB and 4-1BBL. (D,E) The frequency of AMs and IMs in CD45+ cells from the lungs. (F,G) The percentage of IMs expressing 4-1BB and 4-1BBL. The experiments were performed twice with similar results. Data were shown as mean ± SEM (* p ≤ 0.05; ** p ≤ 0.01; *** p ≤ 0.001; ns, not significant).

    Article Snippet: For 4-1BB Fusion Protein Treatment100 or 50 µg of 4-1BB fusion protein (4-1BBIg; cat: 50811-M02H, Sino Biological Inc., Beijing, China) per mouse or 100 µg isotype IgG (human IgG1; Sino Biological Inc.) were injected intraperitoneally into mice, on days 1 and 4 after CS administration (3–4 mice per group).

    Techniques: Expressing, Mouse Assay, Affinity Magnetic Separation