Deficient antiviral immune responses in childhood: Distinct roles of atopy and asthma Simonetta Baraldo, PhD, Marco Contoli, MD, PhD, Erica Bazzan, PhD,

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Deficient antiviral immune responses in childhood: Distinct roles of atopy and asthma Simonetta Baraldo, PhD, Marco Contoli, MD, PhD, Erica Bazzan, PhD, Graziella Turato, PhD, Anna Padovani, BSc, Brunilda Marku, MD, Fiorella Calabrese, MD, Gaetano Caramori, MD, PhD, Andrea Ballarin, MD, Deborah Snijders, MD, Angelo Barbato, MD, Marina Saetta, MD, Alberto Papi, MD Journal of Allergy and Clinical Immunology Volume 130, Issue 6, Pages 1307-1314 (December 2012) DOI: 10.1016/j.jaci.2012.08.005 Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 1 Interferon induction and rhinovirus vRNA levels. A, RV16-induced IFN-β mRNA levels at 8 hours (P = .02, Kruskal-Wallis test). B, RV16-induced IFN-λ mRNA levels at 8 hours (P = .009, Kruskal-Wallis test). C, RV16-induced IFN-β protein levels at 48 hours (P = .02, Kruskal-Wallis test). D, Rhinovirus vRNA levels at 8 hours (P = .02, Kruskal-Wallis test). Bottom and top of the box plot: 25th and 75th percentiles; solid line, median; brackets, 10th and 90th percentiles. Fig 1, A, B, and D: Atopic asthmatic children, n = 8; nonatopic asthmatic children, n = 9; atopic nonasthmatic children, n = 8; control children, n = 9. Fig 1, C: Atopic asthmatic children, n = 6; nonatopic asthmatic children, n = 5; atopic nonasthmatic children, n = 5; control children, n = 5. Journal of Allergy and Clinical Immunology 2012 130, 1307-1314DOI: (10.1016/j.jaci.2012.08.005) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 2 Immunopathologic profile. A, Airway biopsy IL-4 positivity (P = .02, Kruskal-Wallis test). B, Peripheral blood eosinophil counts (P = .002, Kruskal-Wallis test). C, Airway biopsy eosinophil counts (P = .08, Kruskal-Wallis test). D, Epithelial damage (P = .04, Kruskal-Wallis test). Fig 2, A, C, and D: Atopic asthmatic children, n = 6; nonatopic asthmatic children, n = 7; atopic nonasthmatic children, n = 6; control children, n = 6. Fig 2, B: Atopic asthmatic children, n = 8; nonatopic asthmatic children, n = 9; atopic nonasthmatic children, n = 8; control children, n = 9. Journal of Allergy and Clinical Immunology 2012 130, 1307-1314DOI: (10.1016/j.jaci.2012.08.005) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 3 Representative microphotographs of pathologic changes in biopsy specimens related to the impaired innate responses observed ex vivo. A-C, Eosinophils as detected by means of immunostaining with the mAb anti-EG2 (eosinophils are stained in red). D-F, IL-4 positivity as detected by means of immunostaining with the mAb anti–IL-4 (positive cells are stained in red). G-I, Damage to the airway epithelium in sections stained with hematoxylin and eosin. Scale bars = 10 μm. Journal of Allergy and Clinical Immunology 2012 130, 1307-1314DOI: (10.1016/j.jaci.2012.08.005) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 4 Association between immunopathologic profile and interferon production. A and B, RV16-induced IFN-β mRNA (Fig 4, A) and IFN-λ mRNA (Fig 4, B) levels at 8 hours stratified according to eosinophil (eos) levels in airway biopsy specimens. C, Correlation between RV16-dependent induction of IFN-β protein at 48 hours and IL-4 expression (Spearman rank correlation: P = .02, r = −0.58). D, Correlation between RV16-dependent induction of IFN-β protein at 48 hours and degree of epithelial damage (Spearman rank correlation: P = .02, r = −0.55). Journal of Allergy and Clinical Immunology 2012 130, 1307-1314DOI: (10.1016/j.jaci.2012.08.005) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig 5 A, Correlation between total serum IgE levels and RV16-dependent induction of IFN-λ mRNA at 8 hours (Spearman rank correlation: P < .05, r = −0.41). B, Correlation between total serum IgE levels and RV16-dependent induction of vRNA levels at 8 hours (Spearman rank correlation: P < .05, r = 0.44). Journal of Allergy and Clinical Immunology 2012 130, 1307-1314DOI: (10.1016/j.jaci.2012.08.005) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig E1 Journal of Allergy and Clinical Immunology 2012 130, 1307-1314DOI: (10.1016/j.jaci.2012.08.005) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig E2 Journal of Allergy and Clinical Immunology 2012 130, 1307-1314DOI: (10.1016/j.jaci.2012.08.005) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig E3 Journal of Allergy and Clinical Immunology 2012 130, 1307-1314DOI: (10.1016/j.jaci.2012.08.005) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions

Fig E4 Journal of Allergy and Clinical Immunology 2012 130, 1307-1314DOI: (10.1016/j.jaci.2012.08.005) Copyright © 2012 American Academy of Allergy, Asthma & Immunology Terms and Conditions