Context.-Surgical and pathologic handling of lung physically affects lung tissue. This leads to artifacts that alter the morphologic appearance of pulmonary parenchyma.Objective.-To describe and illustrate mechanisms of ex vivo artifacts that may lead to diagnostic pitfalls.Design.-In this study 4 mechanisms of ex vivo artifacts and corresponding diagnostic pitfalls are described and illustrated.Results.-The 4 patterns of artifacts are: (1) surgical collapse, due to the removal of air and blood from pulmonary resections; (2) ex vivo contraction of bronchial and bronchiolar smooth muscle; (3) clamping edema of open lung biopsies; and (4) spreading of tissue fragments and individual cells through a knife surface. Morphologic pitfalls include diagnostic patterns of adenocarcinoma, asthma, constrictive bronchiolitis, and lymphedema.Conclusion.-Four patterns of pulmonary ex vivo artifacts are important to recognize in order to avoid morphologic misinterpretations.

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. @ERSpublications Transcriptome sequencing shows processes in the airways of asthmatics are differently regulated at transcriptomic level

Oral prednisolone changes the transcriptomic profile of the ASM layer in asthma. This indicates that in parallel to antiinflammatory properties, glucocorticoids also exert effects on gene expression of ASM, which is correlated with improved airway function.

In asthma, ECM in ASM is related to the dynamics of airway function in the absence of differences in ECM expression between asthma and controls. This indicates that the ASM layer in its full composition is a major structural component in determining variable airways obstruction in asthma.

Airway smooth muscle from asthma patients can be distinguished from that of atopic and nonatopic control subjects by a specific gene expression profile, which is associated with airway hyper-responsiveness.

Background Airway inflammation in asthma involves innate immune responses. Toll-like receptors (TLRs) and thymic stromal lymphopoietin (TSLP) are thought to be involved in airway inflammation, but their expression in asthmatics’ both large and small airways has not been investigated. Objective To analyze the expression of TLR2, TLR3, TLR4 and TSLP in large and small airways of asthmatics and compare their expression in smoking and nonsmoking asthmatics; to investigate whether TLR expression is associated with eosinophilic or neutrophilic airway inflammation and with Mycoplasma pneumoniae and Chlamydophila pneumoniae infection. Methods Using immunohistochemistry and image analysis, we investigated TLR2, TLR3, TLR4 and TSLP expression in large and small airways of 24 victims of fatal asthma, FA, (13 nonsmokers, 11 smokers) and 9 deceased control subjects (DCtrl). TLRs were also measured in 18 mild asthmatics (MA) and 12 healthy controls (HCtrl). Mycoplasma pneumoniae and Chlamydophila pneumoniae in autopsy lung tissue was analyzed using real-time polymerase chain reaction. Airway eosinophils and neutrophils were measured in all subjects. Results Fatal asthma patients had higher TLR2 in the epithelial and outer layers of large and small airways compared with DCtrls. Smoking asthmatics had lower TLR2 levels in the inner and outer layers of the small airways than nonsmoking asthmatics. TSLP was increased in the epithelial and outer layers of the large airways of FA. FA patients had greater TLR3 expression in the outer layer of large airways and greater TLR4 expression in the outer layer of small airways. Eosinophilic airway inflammation was associated with TLR expression in the epithelium of FA. No bacterial DNA was detected in FA or DCtrls. MA and HCtrls had only a small difference in TLR3 expression. Conclusions and Clinical Relevance Increased expression of TLR 2, 3 and 4 and TSLP in fatal asthma may contribute to the acute inflammation surrounding asthma deaths.

BackgroundAirway remodelling is a feature of asthma including fragmentation of elastic fibres observed in the superficial elastin network of the airway wall. Fibered confocal fluorescence microscopy (FCFM) is a new and non-invasive imaging technique performed during bronchoscopy that may visualize elastic fibres, as shown by in vitro spectral analysis of elastin powder. We hypothesized that FCFM images capture in vivo elastic fibre patterns within the airway wall and that such patterns correspond with airway histology. We aimed to establish the concordance between the bronchial elastic fibre pattern in histology and FCFM. Second, we examined whether elastic fibre patterns in histology and FCFM were different between asthmatic subjects and healthy controls. Finally, the association between these patterns and lung function parameters was investigated.MethodsIn a cross-sectional study comprising 16 subjects (8 atopic asthmatic patients with controlled disease and 8 healthy controls) spirometry and bronchoscopy were performed, with recording of FCFM images followed by endobronchial biopsy at the airway main carina. Elastic fibre patterns in histological sections and FCFM images were scored semi-quantitatively. Agreement between histology and FCFM was analysed using linearly weighted kappa κw.ResultsThe patterns observed in histological sections and FCFM images could be divided into 3 distinct groups. There was good agreement between elastic fibre patterns in histology and FCFM patterns (κw 0.744). The semi-quantitative pattern scores were not different between asthmatic patients and controls. Notably, there was a significant difference in post-bronchodilator FEV1 %predicted between the different patterns by histology (p = 0.001) and FCFM (p = 0.048), regardless of asthma or atopy.ConclusionFCFM captures the elastic fibre pattern within the airway wall in humans in vivo. The association between post-bronchodilator FEV1 %predicted and both histological and FCFM elastic fibre patterns points towards a structure-function relationship between extracellular matrix in the airway wall and lung function.Trial registrationNetherlands Trial Register NTR1306

The complex biology of respiratory diseases such as asthma is feeding the discovery of various disease phenotypes. Although the clinical management of asthma phenotypes by using a single biomarker (e.g., sputum eosinophils) is successful, emerging evidence shows the requirement of multiscale, high-dimensional biological and clinical measurements to capture the complexity of various asthma phenotypes. High-throughput "omics" technologies, including transcriptomics, proteomics, lipidomics, and metabolomics, are increasingly standardized for biomarker discovery in asthma. The leading principle is obeying available guidelines on omics analysis, thereby strictly limiting false discovery. In this review we address the concept of transcriptomics using microarrays or next-generation RNA sequencing and their applications in asthma, highlighting the strengths and limitations of both techniques, and review metabolomics in exhaled air (breathomics) as a noninvasive alternative for sampling the airways directly. These developments will inevitably lead to the integration of molecular signatures in the phenotyping of asthma and other diseases.