1. Investigación

Allergic asthma is a prominent disease especially during childhood. Indoor allergens, in general, and particularly house dust mites (HDM) are the most prevalent sensitizers associated with allergic asthma. Available data show that 65–130 million people are mite-sensitized world-wide and as many as 50% of these are asthmatic. In fact, sensitization to HDM in the first years of life can produce devastating effects on pulmonary function leading to asthmatic syndromes that can be fatal. To date, there has been considerable research into the pathological pathways and structural changes associated with allergic asthma. However, limitations related to the disease heterogeneity and a lack of knowledge into its pathophysiology have impeded the generation of valuable data needed to appropriately phenotype patients and, subsequently, treat this disease. Here, we report a systematic and integral analysis of the disease, from airway remodelling to the immune response taking place throughout the disease stages. We present an overview of metabolomics, the management of complex multifactorial diseases through the analysis of all possible metabolites in a biological sample, obtaining a global interpretation of biological systems. Special interest is placed on the challenges to obtain biological samples and the methodological aspects to acquire relevant information, focusing on the identification of novel biomarkers associated with specific phenotypes of allergic asthma. We also present an overview of the metabolites cited in the literature, which have been related to inflammation and immune response in asthma and other allergy-related diseases.

El asma puede presentar múltiples fenotipos, como el asma alérgico o no alérgico. Su tratamiento es complejo, existiendo pacientes graves que no responden a las medicaciones actualmente disponibles, sufren exacerbaciones frecuentes y presentan comorbilidades como la poliposis nasosinusal. La estratificación de pacientes mediante el uso de biomarcadores permitiría mejorar el tratamiento y descubrir nuevas dianas terapéuticas. Para identificar biomarcadores de estratificación por gravedad, se estudiaron pacientes asmáticos alérgicos estratificados por gravedad utilizando metabolómica y proteómica. Los pacientes con asma grave no controlado presentaron una activación característica de las rutas del ácido araquidónico, la fosfolipasa A2, y la respuesta Th2. Además, para evaluar la contribución del fenotipo alérgico al asma, se realizó un análisis metabolómico de pacientes graves no controlados con y sin alergia. Los pacientes asmáticos alérgicos mostraron una activación de la ruta de la fosfolipasa A2 y una alteración en el perfil de ácidos biliares. Por último, para conocer el papel sistémico y local de la alergia en la poliposis nasal, se estudiaron pacientes con poliposis con y sin alergia utilizando metabolómica de suero y tejido y análisis histológicos. Los pacientes alérgicos presentaron niveles reducidos de lisofosfolípidos, bilirrubina y cortisol; y un mayor número de eosinófilos en sus pólipos.

Background: Asthma is a complex, multifactorial disease often linked with sensitization to house dust mites (HDM). There is a subset of patients that does not respond to available treatments, who present a higher number of exacerbations and a worse quality of life. To understand the mechanisms of poor asthma control and disease severity, we aim to elucidate the metabolic and immunologic routes underlying this specific phenotype and the associated clinical features. Methods: Eighty-seven patients with a clinical history of asthma were recruited and stratified in 4 groups according to their response to treatment: corticosteroid-controlled (ICS), immunotherapy-controlled (IT), biologicals-controlled (BIO) or uncontrolled (UC). Serum samples were analysed by metabolomics and proteomics; and classifiers were built using machine-learning algorithms. Results: Metabolomic analysis showed that ICS and UC groups cluster separately from one another and display the highest number of significantly different metabolites among all comparisons. Metabolite identification and pathway enrichment analysis highlighted increased levels of lysophospholipids related to inflammatory pathways in the UC patients. Likewise, 8 proteins were either upregulated (CCL13, ARG1, IL15 and TNFRSF12A) or downregulated (sCD4, CCL19 and IFNγ) in UC patients compared to ICS, suggesting a significant activation of T cells in these patients. Finally, the machine-learning model built including metabolomic and clinical data was able to classify the patients with an 87.5% accuracy. Conclusions: UC patients display a unique fingerprint characterized by inflammatory-related metabolites and proteins, suggesting a pro-inflammatory environment. Moreover, the integration of clinical and experimental data led to a deeper understanding of the mechanisms underlying UC phenotype.

Metabolomics, understood as the science that manages the study of compounds from the metabolism, is an essential tool for deciphering metabolic changes in disease. The experiments rely on the use of high-throughput analytical techniques such as liquid chromatography coupled to mass spectrometry (LC-ToF MS). This hyphenation has brought positive aspects such as higher sensitivity, specificity and the extension of the metabolome coverage in a single run. The analysis of a high number of samples in a single batch is currently not always feasible due to technical and practical issues (i.e., a drop of the MS signal) which result in the MS stopping during the experiment obtaining more than a single sample batch. In this situation, careful data treatment is required to enable an accurate joint analysis of multi-batch data sets. This paper summarizes the analytical strategies in large-scale metabolomic experiments; special attention has been given to QC preparation troubleshooting and data treatment. Moreover, labeled internal standards analysis and their aim in data treatment, and data normalization procedures (intra- and inter-batch) are described. These concepts are exemplified using a cohort of 165 patients from a study in asthma.

Summary Background: Many epidemiological studies have assessed the prevalence of respiratory allergic disorders in confined geographical locations. However, no study has yet established nationally prevalence data in a uniform manner representing whole countries and, thus, enabling cross-national comparisons. Methods: In 10 European countries, screening of random, representative samples of telephone numbers identified the target population aged 16–60. The inclusion criteria were a positive reporting of respiratory allergy to named allergens and, concomitantly, an unassisted description of appropriate symptoms. To obtain a truly representative, national prevalence of each country, the data were weighted against the actual sex and age composition. Results: 31,065 screening interviews were performed. The nationally balanced prevalence varied significantly among the 10 countries (Po0:001) from 11.7% in Spain to 33.6% in Italy. The overall weighted prevalence for Europe was 24.4%. Comparing males and females, overall, the odds-ratio was 0.874 (Po0:001). For age intervals of 16–29, 30–49, and 50–60 years, the odds-ratios for males were 1.104 (Po0:088), 0.827 (Po0:001), and 0.658 (Po0:001), respectively. The prevalence correlated inversely with age. Conclusions: Respiratory allergic disorders constitute a huge health problem in Europe, and the impact may be increasing as the prevalence is highest among young people.

The aim of this study was to assess the degree of asthma control according to GINA criteria during two different seasons in Spain. An multicenter, longitudinal, epidemiological study with the participation of a sample of physicians in Spain was conducted. Consecutive asthma patients, 18 years of age and older, seeking primary and specialist care were included in the study. Patients were seen during the winter and spring 2004 and were asked about asthma control according to GINA control criteria (daytime and nighttime symptoms, asthma exacerbations, limitations of physical activity, and visits to the emergency department) during the 4 weeks prior to the visit. Control was defined according to daytime and nighttime symptoms. A total of 614 patients participated in the study. The proportion of patients reporting daytime symptoms ‘‘every day’’ or ‘‘most days’’ during the winter versus spring was 40.1% vs. 23% ðPo0:01Þ; 26.9% vs. 14.1% presented symptoms at night ðPo0:01Þ; 11.5% vs. 8.3% had severe exacerbations; 33.5% vs. 35.7% presented symptoms accompanying exercise, and 9.4% vs. 4.3% ðPo0:01Þ had required emergency visits. The number of patients with inadequate control was slightly higher in winter than in spring (74.4% vs. 71%) ðPo0:01Þ. The most commonly prescribed treatment was ICS plus LABAs for both periods.

Eosinophils are important effector cells in allergic inflammation described in allergic rhinitis (AR) and allergic bronchial asthma (BA). During the pollen season serum levels of eosinophil cationic protein (ECP) and eosinophil X protein/eosinophil-derived neurotoxin (EPX/EDN) are increased in BA. The aim of the present study was to evaluate the serum levels of ECP and EPX in pollen atopic patients with AR and BA during the winter. 92 patients were studied. They were divided into three groups: I 29 patients with AR, II 51 patients with BA and 111 12 healthy subjects. Allergic rhinitis and bronchial asthma were diagnosed by routine clinical tests: clinical history, skin tests, total lgE and specific lgE. In addition ECP and EPX were determined in serum. All patients were asymptomatic, stable and without medical treatment. Methacholine challenge test (MCT) was performed in all patients. MCT were positive in 4 patients of group I and 45 patients of group 11. ECP levels (ug/I) were: 21'(1), 24 (II) and 7 (Ill). EPX levels (ug/1) were 35 (I), 45 (II) and 21 (Il l). Statistical differences (p< 0.01) were obseNed both in ECP and EPX levels in patients with MCT positive in relation to patients with MCT negative, and in allergic patients (I and II) in comparison with the healthy subjects (Ill) (p< 0.01). ECP and EPX serum levels are increased in patients with a positive MCT in the winter, out of the pollen season, when patients are asymptomatic, stable and without treatment. This fact suggests that eosinophils play an important role in the pathogenesis of bronchial asthma.