Comparative proteomic analysis identifies specific signature proteins in small extracellular vesicles from granulocytes and CD4+ T cell subpopulations

Proteomic analysis of small extracellular vesicles (sEV) is a powerful tool for the characterization of cell-to-cell signalling. While several studies describe the proteome profile of different immune cell types, none have analysed the content of their delivered sEV. The aim of the present study was to compare the protein signature of sEV delivered from cell subpopulations with importance for chronic inflammatory diseases: granulocytes (i.e., neutrophils and eosinophils) and CD4+ T cells (i.e., TH1, TH2 and TH17). Qualitative (DDA) and quantitative (DIA-SWATH) proteomic analysis of in vitro produced sEV resulted in the identification of proteomic differences depending on the originating cell source. The main differences were found between granulocytes and TH cell-derived sEV, with higher abundance of antimicrobial proteins (e.g., LCN2, LTF, MPO) in sEV from granulocytes and an enrichment of ribosomal proteins (RPL and RPS proteins) in the TH ones. Additionally, we found differentially abundant proteins between neutrophil and eosinophil sEV (e.g., ILF2, LTF, LCN2) and among TH subsets (e.g., ISG15, ITGA4, ITGB2 or NAMPT). These results highlight the potential of sEV proteins as a source of biomarkers for diseases where the different immune cell subsets studied participate, particularly chronic inflammatory pathologies such as asthma or COPD.