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Jethe O. F. Nunes, PhD, Juliana de Souza Apostolico, MSc, David A. G

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Presentation on theme: "Jethe O. F. Nunes, PhD, Juliana de Souza Apostolico, MSc, David A. G"— Presentation transcript:

1 Sleep deprivation predisposes allergic mice to neutrophilic lung inflammation 
Jethe O.F. Nunes, PhD, Juliana de Souza Apostolico, MSc, David A.G. Andrade, PhD, Francieli S. Ruiz, PhD, Edgar R. Fernandes, BSc, Monica L. Andersen, PhD, Alexandre C. Keller, PhD, Daniela S. Rosa, PhD  Journal of Allergy and Clinical Immunology  Volume 141, Issue 3, Pages e4 (March 2018) DOI: /j.jaci Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

2 Fig 1 SD aggravates allergen-induced airway inflammation. A, Experimental protocol. i.n., Intranasal; s.c., subcutaneous. B, Total BALF cell counts. C-E, Mononuclear cell (Fig 1, C), neutrophil (Fig 1, D), and eosinophil (Fig 1, E) counts of cytospin preparations from BALF cells. F-I, Frequency of neutrophils (Fig 1, F), eosinophils (Fig 1, G), mast cells (Fig 1, H), and basophils (Fig 1, I) in the lung determined by using flow cytometry. Data were from 3 independent experiments (n = 5-8 mice per group). Only significant differences between the HS and SD allergic groups are shown. ***P < .001 and ****P < .0001. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

3 Fig 2 SD induces a proinflammatory TH17 profile. A, TNF-α, IL-4, IL-6, and IL-17 levels in culture supernatant from lungs, CLNs, and spleen cells were assessed by using a Cytometric Bead Array. B, IL-4, IL-5, and IL-13 levels in serum were assessed by ELISA. Data were from 3 independent experiments (n = 5-8 mice per group). Only significant differences between the HS and SD allergic groups are shown. **P < .01, ***P < .001, and ****P <  ND, Not detected. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

4 Fig 3 SD leads to allergen-induced lung neutrophilic inflammation in an IL-17 dependent manner. A, Percentage of IL-17+ and TNF-α+ cells in lung CD4+ T cells. Boolean combinations show the frequency of each response based on all possible combinations of cytokine-producing CD4+ T cells. B, Percentage of IL-6+ and TNF-α+ cells in lung macrophages. Boolean combinations show the frequency of each response based on all possible combinations of cytokine-producing macrophages. C, Adoptive transfer scheme. i.n., Intranasal; s.c., subcutaneous. D, Total BALF cell counts. E, Neutrophil counts of cytospin preparations from BALF cells. F, Frequency of neutrophils in the lungs by flow cytometry. Data were from 2 independent experiments (n = 5 mice per group). Only significant differences between the HS and SD allergic groups are shown. *P < .05, **P < .01, and ***P < .001. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

5 Fig 4 SD aggravates allergen-induced lung inflammation in an IL-17RA–dependent manner. A, Total BALF cell counts. B and C, Eosinophil (Fig 4, B) and neutrophil (Fig 4, C) counts of cytospin preparations from BALF cells. D and E, Frequency of eosinophils (Fig 4, D) and neutrophils (Fig 4, E) in the lungs determined by using flow cytometry. Data were from 2 independent experiments (n = 6 mice per group). Only significant differences between the HS and SD allergic groups are shown. **P < .01 and ***P < .001. White bars, WT mice; black bars, IL-17RA–deficient mice (IL-17RA−/−). Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

6 Fig 5 SD induces corticosteroid resistance in allergic mice. A, Total BALF cell counts. B and C, Eosinophil (Fig 5, B) and neutrophil (Fig 5, C) counts of cytospin preparations from BALF cells. D-F, IL-17 (Fig 5, D), TNF-α (Fig 5, E), and IL-6 (Fig 5, F) levels in culture supernatants from lung cells were assessed by using the Cytometric Bead Array. G, Representative lung section stained with hematoxylin and eosin for analysis of cellular inflammation (original magnification ×200). Data were from 2 independent experiments (n = 6 mice per group). DEXA, Dexamethasone. Only significant differences between the vehicle and dexamethasone treated groups are shown. *P < .05, **P < .01, and ∗∗∗∗P < White bars, Vehicle-treated mice; black bars, dexamethasone-treated mice. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

7 Fig E1 Representative dot plots of a multicolor flow cytometry panel used to identify myeloid cell subsets. Mouse lungs were enzymatically digested, and cells were isolated as described in the “Methods” section. After doublet exclusion, immune cells were identified by CD45 staining. Within the CD45+ population, mast cells were gated as c-Kit+FcεRI+ double positive, basophils as c-Kit−CD49+FcεRI+, neutrophils as Gr1+CD11b+, and eosinophils as Gr1lowSSChiIgE+FcεRI+. Comparison of lung cells was done in allergic mice with HS and allergic mice with SD. The Gr1lowSSChi (gate 1) cells were further analyzed for IgE and FcεRI expression, and Gr1hiSSCint cells (gate 2) were further analyzed for CD11b expression. FSC, Forward scatter; SSC, side scatter. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

8 Fig E2 SD did not influence the OVA-specific humoral response. OVA-specific IgG1 (A) and IgE (B) levels were assessed by ELISA. Data were from 3 independent experiments (n = 6 mice per group). Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

9 Fig E3 SD aggravates allergen-induced airway inflammation. A, Experimental protocol. i.n., Intranasal; s.c., subcutaneous. B, Total BALF cell counts. C-E, Differential counts of cytospin preparations from BALF cells. F-I, Frequency of myeloid cell subsets in the lung, as determined by using flow cytometry. J, TNF-α, IL-4, IL-6, and IL-17 levels in culture supernatants from lungs cells were assessed by using a Cytometric Bead Array. Data were from 3 independent experiments (n = 5-8 mice per group). Only significant differences between the HS and SD groups and between the HS and SD allergic groups are shown. **P < .01, ***P < .001, and ∗∗∗∗P <.0001. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

10 Fig E4 SD did not influence corticosterone levels. Corticosterone levels in plasma were assessed by using liquid chromatography tandem mass spectrometry 24 hours after the last set of OVA challenges. Journal of Allergy and Clinical Immunology  , e4DOI: ( /j.jaci ) Copyright © 2017 American Academy of Allergy, Asthma & Immunology Terms and Conditions

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