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The resolution of inflammation is a complex process that is critical for removing inflammatory cells and restoring tissue function. The dysregulation of these mechanisms leads to chronic inflammatory disorders. Platelets, essential cells for preserving homeostasis, are thought to play a role in inflammation as they are a source of immunomodulatory factors. Our aim was to identify key metabolites carried by platelet-derived extracellular vesicles (PL-EVs) in a model of allergic inflammation. PL-EVs were isolated by serial ultracentrifugation using platelet-rich plasma samples obtained from platelet apheresis from severely (n = 6) and mildly (n = 6) allergic patients and non-allergic individuals used as controls (n = 8). PL-EVs were analysed by a multiplatform approach using liquid and gas chromatography coupled to mass spectrometry (LC-MS and GC-MS, respectively). PL-EVs obtained from severely and mildly allergic patients and control individuals presented comparable particle concentrations and sizes with similar protein concentrations. Strikingly, PL-EVs differed in their lipid and metabolic content according to the severity of inflammation. L-carnitine, ceramide (Cer (d18:0/24:0)), and several triglycerides, all of which seem to be involved in apoptosis and regulatory T functions, were higher in PL-EVs from patients with mild allergic inflammation than in those with severe inflammation. In contrast, PL-EVs obtained from patients with severe allergic inflammation showed an alteration in the arachidonic acid pathway. This study demonstrates that PL-EVs carry specific lipids and metabolites according to the degree of inflammation in allergic patients and propose novel perspectives for characterising the progression of allergic inflammation. [ABSTRACT FROM AUTHOR]

The transition from mild to severe allergic phenotypes is still poorly understood and there is an urgent need of incorporating new therapies, accompanied by personalized diagnosis approaches. This work presents the development of a novel targeted metabolomic methodology for the analysis of 36 metabolites related to allergic inflammation, including mostly sphingolipids, lysophospholipids, amino acids, and those of energy metabolism previously identified in non-targeted studies. The methodology consisted of two complementary chromatography methods, HILIC and reversed-phase. These were developed using liquid chromatography, coupled to triple quadrupole mass spectrometry (LC-QqQ-MS) in dynamic multiple reaction monitoring (dMRM) acquisition mode and were validated using ICH guidelines. Serum samples from two clinical models of allergic asthma patients were used for method application, which were as follows: (1) corticosteroid-controlled (ICS, n = 6) versus uncontrolled (UC, n = 4) patients, and immunotherapy-controlled (IT, n = 23) versus biologicals-controlled (BIO, n = 12) patients. The results showed significant differences mainly in lysophospholipids using univariate analyses in both models. Multivariate analysis for model 1 was able to distinguish both groups, while for model 2, the results showed the correct classification of all BIO samples within their group. Thus, this methodology can be of great importance for further understanding the role of these metabolites in allergic diseases as potential biomarkers for disease severity and for predicting patient treatment response. [ABSTRACT FROM AUTHOR]

Thus, metabolomics is an emerging approach that may provide more insight into asthma pathophysiological mechanisms, enable the identification of early biomarkers and targeted personalized therapies, thus reducing the disease burden and societal cost. The inflammatory phenotypes identified to date include eosinophilic or T2 or T2-high asthma, and non-eosinophilic, non-T2 or T2-low asthma, which comprises, in turn, the neutrophilic and paucigranulocytic phenotypes. Asthma is a major non-communicable disease characterized by recurrent attacks of breathlessness and wheezing. Identification of biomarkers and new causative factors may possibly lead to more personalized and precise pathway-specific approaches for asthma diagnosis and treatment. [Extracted from the article]

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. [ABSTRACT FROM AUTHOR]

Emerging evidence suggests that platelets, cytoplasmic fragments derived from megakaryocytes, can no longer be considered just as mediators in hemostasis and coagulation processes, but as key modulators of immunity. Platelets have received increasing attention as the emergence of new methodologies has allowed the characterization of their components and functions in the immune continuum. Platelet activation in infectious and allergic lung diseases has been well documented and associated with bacterial infections reproduced in several animal models of pulmonary bacterial infections. Direct interactions between platelets and bacteria have been associated with increased pulmonary platelet accumulation, whereas bacterial-derived toxins have also been reported to modulate platelet function. Recently, platelets have been found extravascular in the lungs of patients with asthma, and in animal models of allergic lung inflammation. Their ability to interact with immune and endothelial cells and secrete immune mediators makes them one attractive target for biomarker identification that will help characterize their contribution to lung diseases. Here, we present an original review of the last advances in the platelet field with a focus on the contribution of platelets to respiratory infections and allergic-mediated diseases. [ABSTRACT FROM AUTHOR]