A Comprehensive Evaluation of Nasal and Bronchial Cytokines and Chemokines Following Experimental Rhinovirus Infection in Allergic Asthma: Increased Interferons (IFN-γ and IFN-λ) and Type 2 Inflammation (IL-5 and IL-13).

BACKGROUND: Rhinovirus infection is a major cause of asthma exacerbations. OBJECTIVES: We studied nasal and bronchial mucosal inflammatory responses during experimental rhinovirus-induced asthma exacerbations. METHODS: We used nasosorption on days 0, 2-5 and 7 and bronchosorption at baseline and day 4 to sample mucosal lining fluid to investigate airway mucosal responses to rhinovirus infection in patients with allergic asthma (n=28) and healthy non-atopic controls (n=11), by using a synthetic absorptive matrix and measuring levels of 34 cytokines and chemokines using a sensitive multiplex assay. RESULTS: Following rhinovirus infection asthmatics developed more upper and lower respiratory symptoms and lower peak expiratory flows compared to controls (all P<0.05). Asthmatics also developed higher nasal lining fluid levels of an anti-viral pathway (including IFN-γ, IFN-λ/IL-29, CXCL11/ITAC, CXCL10/IP10 and IL-15) and a type 2 inflammatory pathway (IL-4, IL-5, IL-13, CCL17/TARC, CCL11/eotaxin, CCL26/eotaxin-3) (area under curve day 0-7, all P<0.05). Nasal IL-5 and IL-13 were higher in asthmatics at day 0 (P<0.01) and levels increased by days 3 and 4 (P<0.01). A hierarchical correlation matrix of 24 nasal lining fluid cytokine and chemokine levels over 7days demonstrated expression of distinct interferon-related and type 2 pathways in asthmatics. In asthmatics IFN-γ, CXCL10/IP10, CXCL11/ITAC, IL-15 and IL-5 increased in bronchial lining fluid following viral infection (all P<0.05). CONCLUSIONS: Precision sampling of mucosal lining fluid identifies robust interferon and type 2 responses in the upper and lower airways of asthmatics during an asthma exacerbation. Nasosorption and bronchosorption have potential to define asthma endotypes in stable disease and at exacerbation

Rhinovirus-induced VP1-specific Antibodies are Group-specific and Associated With Severity of Respiratory Symptoms

Background: Rhinoviruses (RVs) are a major cause of common colds and induce exacerbations of asthma and chronic inflammatory lung diseases. Methods: We expressed and purified recombinant RV coat proteins VP1-4, non-structural proteins as well as N-terminal fragments of VP1 from four RV strains (RV14, 16, 89, C) covering the three known RV groups (RV-A, RV-B and RV-C) and measured specific IgG-subclass-, IgA- and IgM-responses by ELISA in subjects with different severities of asthma or without asthma before and after experimental infection with RV16. Findings: Before infection subjects showed IgG1 > IgA > IgM > IgG3 cross-reactivity with N-terminal fragments from the representative VP1 proteins of the three RV groups. Antibody levels were higher in the asthmatic group as compared to the non-asthmatic subjects. Six weeks after infection with RV16, IgG1 antibodies showed a group-specific increase towards the N-terminal VP1 fragment, but not towards other capsid and non-structural proteins, which was highest in subjects with severe upper and lower respiratory symptoms. Interpretation: Our results demonstrate that increases of antibodies towards the VP1 N-terminus are group-specific and associated with severity of respiratory symptoms and suggest that it may be possible to develop serological tests for identifying causative RV groups

Type I conventional dendritic cells relate to disease severity in virus-induced asthma exacerbations

RATIONALE: Rhinoviruses are the major precipitant of asthma exacerbations and individuals with asthma experience more severe/prolonged rhinovirus infections. Concurrent viral infection and allergen exposure synergistically increase exacerbation risk. Although dendritic cells orchestrate immune responses to both virus and allergen, little is known about their role in viral asthma exacerbations. OBJECTIVES: To characterize dendritic cell populations present in the lower airways, and to assess whether their numbers are altered in asthma compared to healthy subjects prior to infection and during rhinovirus‐16 infection. METHODS: Moderately‐severe atopic asthmatic patients and healthy controls were experimentally infected with rhinovirus‐16. Bronchoalveolar lavage was collected at baseline, day 3 and day 8 post infection and dendritic cells isolated using fluorescence activated cell sorting. MEASUREMENTS AND MAIN RESULTS: Numbers of type I conventional dendritic cells, which cross prime CD8(+) T helper cells and produce innate interferons, were significantly reduced in the lower airways of asthma patients compared to healthy controls at baseline. This reduction was associated serum IgE at baseline and with reduced numbers of CD8(+) T helper cells and with increased viral replication, airway eosinophils and reduced lung function during infection. IgE receptor expression on lower airway plasmacytoid dendritic cells was significantly increased in asthma, consistent with a reduced capacity to produce innate interferons. CONCLUSIONS: Reduced numbers of anti‐viral type I conventional dendritic cells in asthma are associated with adverse outcomes during rhinovirus infection. This, with increased FcεR1α expression on lower airway plasmacytoid DCs could mediate the more permissive respiratory viral infection observed in asthma patients

IL-8 induction following TLR stimulation in HBECs.

<p>Graphs depict IL-8 production at 8 h following stimulation with Poly I:C [A], CpG-C-ODN [B], CpG-B-ODN [C] and RNA40 [D] and at 24 h following R848 [E] and LPS [F]. Squares represent asthmatics, circles represent non-asthmatics. * = p<0.05, ** = p<0.01, *** = p<0.001.</p

TLR7 stimulation induced type I but not type III IFN in PBMCs.

<p>Graphs depict IFN-α protein production 8 h [A], 24 h [B] and 48 h [C] and IFN-β protein at 8 h [D], 24 h [E] and 48 h [F] post R848 stimulation. Squares represent asthmatics, circles represent non-asthmatics. * = p<0.05, ** = p<0.01, *** = p<0.001.</p

TLR7 stimulation induced type I and III IFN in HBECs.

<p>Graphs depict IFN-β protein [A], IFN-β mRNA [B], IFN-λ protein [C], IFN-λ1 mRNA [D] and IFN-λ2/3 mRNA [E] at 24 h following stimulation with R848. Squares represent asthmatics, circles represent non-asthmatics. * = p<0.05, ** = p<0.01, *** = p<0.001.</p

IL-8 induction following TLR stimulation in PBMCs.

<p>Graphs depict IL-8 production at 8 h following stimulation with PIC [A], R848 [B], LPS [C] and RNA40 [D], 24 h following PIC [E], R848 [F], CpG-C-ODN [G], CpG-B-ODN [H], LPS [I] and RNA40 [J] and 48 h following PIC [K]. Squares represent asthmatics, circles represent non-asthmatics. * = p<0.05, ** = p<0.01, *** = p<0.001.</p

IL-6 induction following TLR stimulation in PBMCs.

<p>Graphs depict IL-6 production at 8 h following stimulation with PIC [A], R848 [D], LPS [G] and LPS [K], 24 h following PIC [B], R848 [E], LPS [H] and LPS [L] and 48 h following PIC [C], R848 [F], LPS [J] and LPS [M]. Squares represent asthmatics, circles represent non-asthmatics. * = p<0.05, ** = p<0.01, *** = p<0.001.</p