CD18-mediated adhesion is required for lung inflammation induced by mononuclear cell-derived extracellular vesicles

Abstract Extracellular vesicles are submicron vesicles that upregulate the synthesis of proinflammatory mediators by lung epithelial cells. We investigated whether these structures adhere to lung epithelial cells, and whether adhesion is a prerequisite for their proinflammatory activity. Extracellular vesicles were generated by stimulation of normal human mononuclear cells with the calcium ionophore A23187, and labelled with carboxyfluorescein diacetate succinimidyl ester. Adhesion of vesicles to monolayers of immortalized bronchial epithelial cells (16HBE) and alveolar cells (A549) was analysed by fluorescence microscopy. The role of candidate adhesion receptors was evaluated with inhibitory monoclonal antibodies and soluble peptides. The synthesis of proinflammatory mediators was assessed by ELISA. Transmission electron microscopy confirmed the generation of closed vesicles with an approximate size range between 50 and 600 nm. Adhesion of extracellular vesicles to epithelial cells was minimal in baseline conditions and was upregulated upon stimulation of the latter with tumour necrosis factor-α. Adhesion was blocked by an anti-CD18 antibody and by peptides containing the sequence RGD. The same molecules also blocked the upregulation of the synthesis of interleukin-8 and monocyte chemotactic protein-1 induced by extracellular vesicles. Summary statementExtracellular vesicles upregulate the synthesis of proinflammatory mediators by lung epithelial cells. CD18-mediated adhesion to target cells is required for this proinflammatory effect and might represent a target for anti-inflammatory therapy.