Proteomic profiling of urinary small extracellular vesicles in children with pneumonia: a pilot study

Extracellular Vesicles

Background Small extracellular vesicles (sEV) play a crucial role in immune responses to viral infection. However, the composition of sEV derived from children with viral pneumonia remains ill defined. Methods First, we performed mass spectrometry-based label-free proteomic analysis of urinary sEV in 7 children with viral pneumonia, 4 children with Mycoplasma pneumoniae pneumonia and 20 healthy children. Then a total of 33 proteins were selected to validate by multiple reaction monitoring analysis in an independent cohort of 20 healthy children and 29 children with pneumonia. Results In the discovery phase, a total of 1621 proteins were identified, while 260 proteins have differential expression in children with viral pneumonia compared to healthy children. Biological pathways primarily associated with neutrophil degranulation, carbohydrate metabolism and endocytosis were enriched in children with viral pneumonia. Finally, the abundance of eight proteins was verified to be significantly higher in children with viral pneumonia than in healthy children. Conclusions This pilot study with proteomic profiles of urinary sEV provided insights to the host response to viral pathogen exposure and potential diagnostic biomarkers for children with viral pneumonia, and served as the basis for understanding the fundamental biology of infection.

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Recent Publications

Cigarette smoke (CS) represents one of the most relevant environmental risk factors for several chronic pathologies. Tissue damage caused by CS exposure is mediated, at least in part, by oxidative stress induced by its toxic and pro-oxidant components. Evidence demonstrates that extracellular vesicles (EVs) released by various cell types exposed to CS extract (CSE) are characterized by altered biochemical cargo and gained pathological properties. In the present study, we evaluated the content of oxidized proteins and phospholipid fatty acid profiles of EVs released by human bronchial epithelial BEAS-2B cells treated with CSE. This specific molecular characterization has hitherto not been performed. After confirmation that CSE reduces viability of BEAS-2B cells and elevates intracellular ROS levels, in a dose-dependent manner, we demonstrated that 24 h exposure at 1% CSE, a concentration that only slight modifies cell viability but increases ROS levels, was able to increase carbonylated protein levels in cells and released EVs. The release of oxidatively modified proteins via EVs might represent a mechanism used by cells to remove toxic proteins in order to avoid their intracellular overloading. Moreover, 1% CSE induced only few changes in the fatty acid asset in BEAS-2B cell membrane phospholipids, whereas several rearrangements were observed in EVs released by CSE-treated cells. The impact of changes in acyl chain composition of CSE-EVs accounted for the increased saturation levels of phospholipids, a membrane parameter that might influence EV stability, uptake and, at least in part, EV-mediated biological effects. The present in vitro study adds new information concerning the biochemical composition of CSE-related EVs, useful to predict their biological effects on target cells. Furthermore, the information regarding the presence of oxidized proteins and the specific membrane features of CSE-related EVs can be useful to define the utilization of circulating EVs as marker for diagnosing of CS-induced lung damage and/or CS-related diseases.

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