Mast cell-derived nanovesicles potentiate allergic inflammation

Mast cells (MCs) play a major role in allergic diseases: After activation by the high affinity receptor for IgE (FcepsilonRI), they release a high number of pro-inflammatory factors. Moreover, MCs can also produce exosomes, nanosized vesicles that might act as vehicles for intercellular communication. However, evidence of a role for MC-derived exosomes during allergic inflammation is limited. The aim of my PhD project was to characterize exosome-like vesicles released by unstimulated and IgE-stimulated MCs. Secreted nanovesicles were purified from the supernatant of MCs and from human sera by a series of centrifugations and ultracentrifugations and, in order to assess their purity, they were characterized in terms of morphology, size and marker expression. The presence of FcepsilonRI/IgE complexes was evaluated by western blotting combined with FACS analysis and ELISA. MC activation was measured by degranulation assay and multiplex immune-assay, and exosome uptake was analysed by confocal microscopy and flow cytometry. We found that FcepsilonRI engagement increases the amount of exosome release compared to the constitutive production. Moreover, only exosomes secreted upon antigen stimulation display both surface IgE and antigen. We have also demonstrated that vesicles exposing IgE and antigen are efficiently internalized by sensitized MCs and are able to induce their degranulation as well as pro-inflammatory cytokine production. Finally, we found that exosomes purified from serum of atopic patients are also endowed with FcepsilonRI/IgE complexes. All together, these findings reveal a potential novel mechanism of allergic reaction amplification.