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Macrophage-expressed IFN-β contributes to apoptotic alveolar epithelial cell injury in severe influenza virus pneumonia.

Högner K, Wolff T, Pleschka S, Plog S, Gruber AD, Kalinke U, Walmrath HD, Bodner J, Gattenlöhner S, Lewe-Schlosser P, Matrosovich M, Seeger W, Lohmeyer J, Herold S - PLoS Pathog. (2013)

Bottom Line: Bone marrow chimeric mice lacking these signalling mediators in resident and lung-recruited AM and mice subjected to alveolar neutralization of IFN-β and TRAIL displayed reduced alveolar epithelial cell apoptosis and attenuated lung injury during severe IV pneumonia.Together, we demonstrate that macrophage-released type I IFNs, apart from their well-known anti-viral properties, contribute to IV-induced AEC damage and lung injury by autocrine induction of the pro-apoptotic factor TRAIL.Our data suggest that therapeutic targeting of the macrophage IFN-β-TRAIL axis might represent a promising strategy to attenuate IV-induced acute lung injury.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine II, University of Giessen Lung Center, Giessen, Germany.

ABSTRACT
Influenza viruses (IV) cause pneumonia in humans with progression to lung failure and fatal outcome. Dysregulated release of cytokines including type I interferons (IFNs) has been attributed a crucial role in immune-mediated pulmonary injury during severe IV infection. Using ex vivo and in vivo IV infection models, we demonstrate that alveolar macrophage (AM)-expressed IFN-β significantly contributes to IV-induced alveolar epithelial cell (AEC) injury by autocrine induction of the pro-apoptotic factor TNF-related apoptosis-inducing ligand (TRAIL). Of note, TRAIL was highly upregulated in and released from AM of patients with pandemic H1N1 IV-induced acute lung injury. Elucidating the cell-specific underlying signalling pathways revealed that IV infection induced IFN-β release in AM in a protein kinase R- (PKR-) and NF-κB-dependent way. Bone marrow chimeric mice lacking these signalling mediators in resident and lung-recruited AM and mice subjected to alveolar neutralization of IFN-β and TRAIL displayed reduced alveolar epithelial cell apoptosis and attenuated lung injury during severe IV pneumonia. Together, we demonstrate that macrophage-released type I IFNs, apart from their well-known anti-viral properties, contribute to IV-induced AEC damage and lung injury by autocrine induction of the pro-apoptotic factor TRAIL. Our data suggest that therapeutic targeting of the macrophage IFN-β-TRAIL axis might represent a promising strategy to attenuate IV-induced acute lung injury.

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IV-induced macrophage IFN-β upregulates macrophage TRAIL.(A) Murine AM were treated ex vivo with rIFN-β at the given concentrations and TRAIL mRNA expression was quantified and is depicted as fold induction of unstimulated controls. (B) Murine AM were A/PR8-infected (MOI = 0.1) or treated with 180 U/ml rIFN-β ex vivo in presence of a protease inhibitor cocktail to prevent TRAIL shedding and mTRAIL abundance was analysed by FACS after 24 h. Shown are histograms from a representative experiment (top panel) or mean fluorescence intensities (MFI, bottom panel). (C) Murine wt AM were A/PR8 infected at the given MOI and treated with anti-IFN-β Ab, Jak/STAT inhibitor, or DMSO/isotype IgG Ab as control. Murine ifnar−/− AM were A/PR8 infected and left untreated. TRAIL mRNA expression was quantified and is depicted as fold induction of mock-infected cells. Bar graphs represent means ± SD of (A) 6; (B) 4 and (C) 5 independent experiments. * p<0.05; ** p<0.01; ***p<0.001; MOI, multiplicity of infection; IgG, IgG isotype control; rIFN-β, recombinant IFN-β; mTRAIL, membrane bound TRAIL; Ab, antibody.
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ppat-1003188-g003: IV-induced macrophage IFN-β upregulates macrophage TRAIL.(A) Murine AM were treated ex vivo with rIFN-β at the given concentrations and TRAIL mRNA expression was quantified and is depicted as fold induction of unstimulated controls. (B) Murine AM were A/PR8-infected (MOI = 0.1) or treated with 180 U/ml rIFN-β ex vivo in presence of a protease inhibitor cocktail to prevent TRAIL shedding and mTRAIL abundance was analysed by FACS after 24 h. Shown are histograms from a representative experiment (top panel) or mean fluorescence intensities (MFI, bottom panel). (C) Murine wt AM were A/PR8 infected at the given MOI and treated with anti-IFN-β Ab, Jak/STAT inhibitor, or DMSO/isotype IgG Ab as control. Murine ifnar−/− AM were A/PR8 infected and left untreated. TRAIL mRNA expression was quantified and is depicted as fold induction of mock-infected cells. Bar graphs represent means ± SD of (A) 6; (B) 4 and (C) 5 independent experiments. * p<0.05; ** p<0.01; ***p<0.001; MOI, multiplicity of infection; IgG, IgG isotype control; rIFN-β, recombinant IFN-β; mTRAIL, membrane bound TRAIL; Ab, antibody.

Mentions: Given the findings that peak TRAIL expression coincided with peak IFN-β expression at 16 h pi in murine AM (Fig. 1E, G) and that TRAIL and IFN-β showed similar IV strain-specific induction patterns in both murine and human AM (Fig. S2), we speculated that macrophage TRAIL expression was dependent on autocrine IFN-β-induced signaling. Indeed, when we stimulated non-infected murine AM with recombinant IFN-β, we detected a time- and dose-dependent increase of TRAIL mRNA (Fig. 3A) and significantly enhanced cell surface TRAIL expression after 24 h of IFN-β stimulation, which was comparable to A/PR8-induced expression (Fig. 3B). Finally, blockade of IFN-β-induced signalling via the type I IFN receptor (IFNAR) nearly abolished A/PR8-induced TRAIL mRNA upregulation in AM regardless of the MOI applied, as demonstrated by either addition of neutralizing anti-IFN-β Ab to the culture medium, use of ifnar−/− AM, or application of an inhibitor of the IFNAR downstream mediators, Jak and STAT (Fig. 3C). Collectively, these data demonstrate that macrophage-released IFN-β induces expression of the pro-apoptotic factor TRAIL in AM upon IV infection in an autocrine fashion.


Macrophage-expressed IFN-β contributes to apoptotic alveolar epithelial cell injury in severe influenza virus pneumonia.

Högner K, Wolff T, Pleschka S, Plog S, Gruber AD, Kalinke U, Walmrath HD, Bodner J, Gattenlöhner S, Lewe-Schlosser P, Matrosovich M, Seeger W, Lohmeyer J, Herold S - PLoS Pathog. (2013)

IV-induced macrophage IFN-β upregulates macrophage TRAIL.(A) Murine AM were treated ex vivo with rIFN-β at the given concentrations and TRAIL mRNA expression was quantified and is depicted as fold induction of unstimulated controls. (B) Murine AM were A/PR8-infected (MOI = 0.1) or treated with 180 U/ml rIFN-β ex vivo in presence of a protease inhibitor cocktail to prevent TRAIL shedding and mTRAIL abundance was analysed by FACS after 24 h. Shown are histograms from a representative experiment (top panel) or mean fluorescence intensities (MFI, bottom panel). (C) Murine wt AM were A/PR8 infected at the given MOI and treated with anti-IFN-β Ab, Jak/STAT inhibitor, or DMSO/isotype IgG Ab as control. Murine ifnar−/− AM were A/PR8 infected and left untreated. TRAIL mRNA expression was quantified and is depicted as fold induction of mock-infected cells. Bar graphs represent means ± SD of (A) 6; (B) 4 and (C) 5 independent experiments. * p<0.05; ** p<0.01; ***p<0.001; MOI, multiplicity of infection; IgG, IgG isotype control; rIFN-β, recombinant IFN-β; mTRAIL, membrane bound TRAIL; Ab, antibody.
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Related In: Results  -  Collection

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ppat-1003188-g003: IV-induced macrophage IFN-β upregulates macrophage TRAIL.(A) Murine AM were treated ex vivo with rIFN-β at the given concentrations and TRAIL mRNA expression was quantified and is depicted as fold induction of unstimulated controls. (B) Murine AM were A/PR8-infected (MOI = 0.1) or treated with 180 U/ml rIFN-β ex vivo in presence of a protease inhibitor cocktail to prevent TRAIL shedding and mTRAIL abundance was analysed by FACS after 24 h. Shown are histograms from a representative experiment (top panel) or mean fluorescence intensities (MFI, bottom panel). (C) Murine wt AM were A/PR8 infected at the given MOI and treated with anti-IFN-β Ab, Jak/STAT inhibitor, or DMSO/isotype IgG Ab as control. Murine ifnar−/− AM were A/PR8 infected and left untreated. TRAIL mRNA expression was quantified and is depicted as fold induction of mock-infected cells. Bar graphs represent means ± SD of (A) 6; (B) 4 and (C) 5 independent experiments. * p<0.05; ** p<0.01; ***p<0.001; MOI, multiplicity of infection; IgG, IgG isotype control; rIFN-β, recombinant IFN-β; mTRAIL, membrane bound TRAIL; Ab, antibody.
Mentions: Given the findings that peak TRAIL expression coincided with peak IFN-β expression at 16 h pi in murine AM (Fig. 1E, G) and that TRAIL and IFN-β showed similar IV strain-specific induction patterns in both murine and human AM (Fig. S2), we speculated that macrophage TRAIL expression was dependent on autocrine IFN-β-induced signaling. Indeed, when we stimulated non-infected murine AM with recombinant IFN-β, we detected a time- and dose-dependent increase of TRAIL mRNA (Fig. 3A) and significantly enhanced cell surface TRAIL expression after 24 h of IFN-β stimulation, which was comparable to A/PR8-induced expression (Fig. 3B). Finally, blockade of IFN-β-induced signalling via the type I IFN receptor (IFNAR) nearly abolished A/PR8-induced TRAIL mRNA upregulation in AM regardless of the MOI applied, as demonstrated by either addition of neutralizing anti-IFN-β Ab to the culture medium, use of ifnar−/− AM, or application of an inhibitor of the IFNAR downstream mediators, Jak and STAT (Fig. 3C). Collectively, these data demonstrate that macrophage-released IFN-β induces expression of the pro-apoptotic factor TRAIL in AM upon IV infection in an autocrine fashion.

Bottom Line: Bone marrow chimeric mice lacking these signalling mediators in resident and lung-recruited AM and mice subjected to alveolar neutralization of IFN-β and TRAIL displayed reduced alveolar epithelial cell apoptosis and attenuated lung injury during severe IV pneumonia.Together, we demonstrate that macrophage-released type I IFNs, apart from their well-known anti-viral properties, contribute to IV-induced AEC damage and lung injury by autocrine induction of the pro-apoptotic factor TRAIL.Our data suggest that therapeutic targeting of the macrophage IFN-β-TRAIL axis might represent a promising strategy to attenuate IV-induced acute lung injury.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine II, University of Giessen Lung Center, Giessen, Germany.

ABSTRACT
Influenza viruses (IV) cause pneumonia in humans with progression to lung failure and fatal outcome. Dysregulated release of cytokines including type I interferons (IFNs) has been attributed a crucial role in immune-mediated pulmonary injury during severe IV infection. Using ex vivo and in vivo IV infection models, we demonstrate that alveolar macrophage (AM)-expressed IFN-β significantly contributes to IV-induced alveolar epithelial cell (AEC) injury by autocrine induction of the pro-apoptotic factor TNF-related apoptosis-inducing ligand (TRAIL). Of note, TRAIL was highly upregulated in and released from AM of patients with pandemic H1N1 IV-induced acute lung injury. Elucidating the cell-specific underlying signalling pathways revealed that IV infection induced IFN-β release in AM in a protein kinase R- (PKR-) and NF-κB-dependent way. Bone marrow chimeric mice lacking these signalling mediators in resident and lung-recruited AM and mice subjected to alveolar neutralization of IFN-β and TRAIL displayed reduced alveolar epithelial cell apoptosis and attenuated lung injury during severe IV pneumonia. Together, we demonstrate that macrophage-released type I IFNs, apart from their well-known anti-viral properties, contribute to IV-induced AEC damage and lung injury by autocrine induction of the pro-apoptotic factor TRAIL. Our data suggest that therapeutic targeting of the macrophage IFN-β-TRAIL axis might represent a promising strategy to attenuate IV-induced acute lung injury.

Show MeSH
Related in: MedlinePlus