Annemiek Dijkhuis

Transcriptome sequencing (RNA-Seq) of human endobronchial biopsies: asthma versus controls

The cellular and molecular pathways in asthma are highly complex. Increased understanding can be obtained by unbiased transcriptomic analysis (RNA-Seq). We hypothesised that the transcriptomic profile of whole human endobronchial biopsies differs between asthma patients and controls. First, we investigated the feasibility of obtaining RNA from whole endobronchial biopsies suitable for RNA-Seq. Secondly, we examined the difference in transcriptomic profiles between asthma and controls. This cross-sectional study compared four steroid-free atopic asthma patients and five healthy nonatopic controls. Total RNA from four biopsies per subject was prepared for RNA-Seq. Comparison of the numbers of reads per gene in asthma and controls was based on the Poisson distribution. 46 genes were differentially expressed between asthma and controls, including pendrin, periostin and BCL2. 10 gene networks were found to be involved in cellular morphology, movement and development. RNA isolated from whole human endobronchial biopsies is suitable for RNA-Seq, showing different transcriptomic profiles between asthma and controls. Novel and confirmative genes were found to be linked to asthma. These results indicate that biological processes in the airways of asthma patients are regulated differently when compared to controls, which may be relevant for the pathogenesis and treatment of the disease. Transcriptome sequencing shows processes in the airways of asthmatics are differently regulated at transcriptomic level http://ow.ly/kDnln

Systemic tryptophan and kynurenine catabolite levels relate to severity of rhinovirus-induced asthma exacerbation: a prospective study with a parallel-group design

Background Patients with allergic asthma have exacerbations which are frequently caused by rhinovirus infection. The antiviral tryptophan-catabolising enzyme indoleamine 2,3-dioxygenase (IDO) is induced by interferon-γ and suppressed by Th2 mediators interleukin (IL)-4 and IL-13. We hypothesised that local IDO activity after viral airway infection is lower in patients with allergic asthma than in healthy controls. Objective To determine whether IDO activity differs between patients with allergic asthma and healthy individuals before and after rhinovirus infection. Methods Healthy individuals and patients with allergic asthma were experimentally infected with low-dose (10 TCID50) rhinovirus 16. Blood, bronchoalveolar lavage fluid and exhaled breath condensate (for mass spectrometry by UPLC-MS/MS) were obtained before and after rhinovirus challenge. Results IDO activity was not induced by rhinovirus infection in either group, despite increases in cold scores. However, baseline pulmonary IDO activity was lower in patients with allergic asthma than in healthy individuals. In contrast, systemic tryptophan and its catabolites were markedly higher in patients with allergic asthma. Moreover, systemic quinolinic acid and tryptophan were associated with eosinophil cationic protein (r=0.43 and r=0.78, respectively) and eosinophils (r=0.38 and r=0.58, respectively) in bronchoalveolar lavage fluid and peak asthma symptom scores after rhinovirus challenge (r=0.53 and r=0.64, respectively). Conclusions Rhinovirus infection by itself induces no IDO activity, but the reduced pulmonary IDO activity in patients with allergic asthma at baseline may underlie a reduced control of viral infections. Notably, the enhanced systemic catabolism of tryptophan in patients with allergic asthma was strongly related to the outcome of rhinovirus challenge in asthma and may serve as a prognostic factor.

An early innate response underlies severe influenza-induced exacerbations of asthma in a novel steroid-insensitive and anti-IL-5-responsive mouse model

Acute worsening of asthma symptoms (exacerbation) is predominantly triggered by respiratory viruses, with influenza causing the most severe exacerbations. The lack of an adequate animal model hampers mechanistic insight and the development of new therapeutics.

Interferon-induced epithelial response to rhinovirus-16 in asthma relates to inflammation and FEV1

Bronchial epithelial cells from mild asthma patients in response to in vivo rhinovirus-16 challenge, display a marked interferon-induced response 6 days post-infection, which correlates with eosinophilic inflammation and virus-induced decline of lung function.

IL-33 drives influenza-induced asthma exacerbations by halting innate and adaptive anti-viral immunity

Background Influenza virus triggers severe asthma exacerbations for which no adequate treatment is available. It is known that IL‐33 levels correlate with exacerbation severity, but its role in the immunopathogenesis of exacerbations has remained elusive. Objective We hypothesized that IL‐33 is necessary to drive asthma exacerbations. We intervened with the IL‐33 cascade and sought to dissect its role, also in synergy with thymic stromal lymphopoietin (TSLP), in airway inflammation, antiviral activity, and lung function. We aimed to unveil the major source of IL‐33 in the airways and IL‐33–dependent mechanisms that underlie severe asthma exacerbations. Methods Patients with mild asthma were experimentally infected with rhinovirus. Mice were chronically exposed to house dust mite extract and then infected with influenza to resemble key features of exacerbations in human subjects. Interventions included the anti–IL‐33 receptor ST2, anti–TSLP, or both. Results We identified bronchial ciliated cells and type II alveolar cells as a major local source of IL‐33 during virus‐driven exacerbation in human subjects and mice, respectively. By blocking ST2, we demonstrated that IL‐33 and not TSLP was necessary to drive exacerbations. IL‐33 enhanced airway hyperresponsiveness and airway inflammation by suppressing innate and adaptive antiviral responses and by instructing epithelial cells and dendritic cells of house dust mite–sensitized mice to dampen IFN‐&bgr; expression and prevent the TH1‐promoting dendritic cell phenotype. IL‐33 also boosted luminal NETosis and halted cytolytic antiviral activities but did not affect the TH2 response. Conclusion Interventions targeting the IL‐33/ST2 axis could prove an effective acute short‐term therapy for virus‐induced asthma exacerbations. Graphical abstract Figure. No Caption available.

IL33 and TSLP involvements in influenza-induced exacerbation in a murine model of chronic asthma

Introduction: Viral airway infections cause acute worsening of asthma symptoms and additional accumulation of inflammatory cell subsets in the airways. Here we aimed to dissect the innate immune response that might underlie virus-induced exacerbations in an adequate murine model of exacerbating asthma. Methods: Mice were sensitized i.n. to HDM extract for 5 days/week for 5 consecutive weeks. On day 29 mice were infected with low dose of influenza A/X31; 3 day before and after the virus infection the animals have been treated with anti TSLP and IL33 receptors (T1/ST2 and TSLPR, respectively) mAbs. The airways cellular influx was assessed, cytokines were measured in BALF, lung function and histology were determined in different time points after infection. Results: Infected sensitized mice infected showed an exacerbation of allergic inflammation as reflected by a prolonged AHR and a boosted inflammatory influx into the airways, in comparison to infected non-sensitized mice. As an early event upon influenza infection, sensitized mice showed the induction of IL-33 and TSLP paralleled by the increased influx into the airways of ILC2, basophils, NK and NKT cells. Blocking the effect of IL-33 and TSLP using anti T1/ST2 and TSLPR mAbs the NKT cells, basophils, ILC2 recruitments into the airways were attenuated; the eosinophilic and neutrophilic influxes were reduced in BALF. Conclusions: Influenza infection of sensitized mice boosted the induction of crucial effectors of the innate immune response, leading the worsening of a broad inflammatory response into the airways. Limiting IL33 and TSLP can dampen airway inflammation and virus-induced changes in HDM-sensitized mice.

Antiviral effect of eosinophils in respiratory tract infection with influenza virus in mice

Rationale Previous studies in an acute house dust mite murine model of allergic asthma have shown that allergic mice have an improved antiviral response to infection with influenza (A/PR/8/34) compared to non-allergic mice as reflected by lower viral titres in the lung and decreased weight loss. These findings demonstrate that allergic mice are able to clear the infection faster. Interestingly, we observed a synergistically enhanced eosinophilic response in bronchoalveolar lavage (BAL) of these mice in response to the viral infection.

Influenza exacerbations in an acute and chronic house dust mite ‘asthma’ model

Methods In the acute asthma model, mice were sensitised intranasally with HDM or PBS on three consecutive days and challenged two weeks later. Two days after the challenge the mice were infected intranasally with 10 TCID50 influenza (A/PR/8/34) or PBS and sacrificed 4, 8 or 14 days after infection. In the chronic asthma model the mice receive influenza in the 5th week of sensitisation (intranasally, 5 days a week for 5 weeks). In both models we assess(ed) cellular influx by analysing bronchoalveolar lavage fluid (BALF), and cytokines and viral load in lung lysate. PenH is determined as a measure of airway hyperresponsiveness (AHR).