Dominika Polak1, Christine Hafner2, Peter Briza3, Claudia Kitzmüller1, Adelheid Elbe-Bürger4, Nazanin Samadi1, Maria Gschwandtner5, Wolfgang Pfützner6, Gerhard J Zlabinger7, Beatrice Jahn-Schmid1, Barbara Bohle8. 1. Department of Pathophysiology and Allergy Research, Center of Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Vienna, Austria. 2. Department of Dermatology, University Hospital St Pölten, Karl Landsteiner University of Health Sciences, St. Pölten, Austria. 3. Department of Molecular Biology, University of Salzburg, Salzburg, Austria. 4. Department of Dermatology, Medical University of Vienna, Vienna, Austria. 5. Department of Dermatology, Research Division of Biology and Pathobiology of the Skin, Medical University of Vienna, Vienna, Austria. 6. Department of Dermatology and Allergology, Philipps University Marburg, Marburg, Germany. 7. Institute of Immunology, Medical University of Vienna, Vienna, Austria. 8. Department of Pathophysiology and Allergy Research, Center of Pathophysiology, Infectiology and Immunology, Medical University of Vienna, Vienna, Austria. Electronic address: barbara.bohle@meduniwien.ac.at.
Abstract
BACKGROUND: Neutrophils and allergen-specific T cells accumulate in patients with allergic late-phase reactions (LPRs). Their presence is associated with severe inflammation. Cytokines, such as GM-CSF, IFN-γ, and IL-3, which are typically found in patients with allergic LPRs, have been proposed to convert neutrophils into antigen-presenting cells (APCs). OBJECTIVE: We sought to assess the antigen-processing and antigen-presenting capacities of neutrophils from allergic patients. METHODS: Neutrophils were isolated from peripheral blood of donors with birch pollen allergy and stimulated with GM-CSF, IFN-γ, and IL-3. The viability and expression of HLA-DR, CD80, and CD86 were assessed by using flow cytometry. HLA-DM expression was analyzed by means of immunoblotting. Allergen uptake was studied after fluorescence labeling of the major birch pollen allergen Bet v 1. Bet v 1 was digested with neutrophilic endolysosomal extracts, and the resulting fragments were sequenced by using mass spectrometry. Neutrophils were used as APCs in coculture experiments with autologous HLA-DR-restricted and Bet v 1-specific T-cell clones reactive with epitopes in different regions of the allergen. In all experiments monocytes were used for comparison. Fluids from suction blisters formed on top of LPRs induced by using intradermal allergen injection were assessed for HLA-DR+ neutrophils by using flow cytometry. RESULTS: The cytokines significantly enhanced the survival, allergen uptake, and expression of HLA-DM and HLA-DR on neutrophils. Neutrophils rapidly degraded Bet v 1 into fragments containing all relevant T-cell epitopes. Cytokine-activated, allergen-pulsed neutrophils induced proliferative and cytokine responses of Bet v 1-specific T cells irrespective of epitope specificity, confirming that they fully processed and presented the allergen. HLA-DR+ neutrophils were detected in patients with cutaneous allergic LPRs. CONCLUSION: Neutrophils can serve as APCs for local allergen-specific effector T cells in patients with allergic LPRs.
BACKGROUND:Neutrophils and allergen-specific T cells accumulate in patients with allergic late-phase reactions (LPRs). Their presence is associated with severeinflammation. Cytokines, such as GM-CSF, IFN-γ, and IL-3, which are typically found in patients with allergic LPRs, have been proposed to convert neutrophils into antigen-presenting cells (APCs). OBJECTIVE: We sought to assess the antigen-processing and antigen-presenting capacities of neutrophils from allergicpatients. METHODS:Neutrophils were isolated from peripheral blood of donors with birch pollen allergy and stimulated with GM-CSF, IFN-γ, and IL-3. The viability and expression of HLA-DR, CD80, and CD86 were assessed by using flow cytometry. HLA-DM expression was analyzed by means of immunoblotting. Allergen uptake was studied after fluorescence labeling of the major birch pollen allergen Bet v 1. Bet v 1 was digested with neutrophilic endolysosomal extracts, and the resulting fragments were sequenced by using mass spectrometry. Neutrophils were used as APCs in coculture experiments with autologous HLA-DR-restricted and Bet v 1-specific T-cell clones reactive with epitopes in different regions of the allergen. In all experiments monocytes were used for comparison. Fluids from suction blisters formed on top of LPRs induced by using intradermal allergen injection were assessed for HLA-DR+ neutrophils by using flow cytometry. RESULTS: The cytokines significantly enhanced the survival, allergen uptake, and expression of HLA-DM and HLA-DR on neutrophils. Neutrophils rapidly degraded Bet v 1 into fragments containing all relevant T-cell epitopes. Cytokine-activated, allergen-pulsed neutrophils induced proliferative and cytokine responses of Bet v 1-specific T cells irrespective of epitope specificity, confirming that they fully processed and presented the allergen. HLA-DR+ neutrophils were detected in patients with cutaneous allergic LPRs. CONCLUSION:Neutrophils can serve as APCs for local allergen-specific effector T cells in patients with allergic LPRs.
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