It is now evident that cell activation is associated with rapid plasma membrane re-arrangements; the production of small particles, often released, would then occur: they have been named microvesicles or ectosomes. Importantly, they are not just by-products but, rather, they seem to display important biological functions. The recent notion that ectosomes from human PMN are anti-inflammatory whereas those obtained from other leukocyte sources are pro-inflammatory [1], prompted us to test whether this dichotomy was associated with presence of the endogenous annexin 1 (AnxA1). This pro-resolution and homeostatic factor induced by glucocorticoids, highly abundant in human PMN: ~60-70% of cell associated AnxA1 contained in subcellular organelles, acts an important brake signal on their activation and trafficking. Ectosomes were produced by PBMC or PMN (50 million per experiment) incubation with HUVEC monolayers in the presence of 1 µM fMLP (60 min at 37°C). Ectosomes were then separated from the cell supernatant by ultracentrifugation and their content analysed by FACS analysis, Western Blotting and Mass Spectrometry (Q-TOF) techniques. The presence of a specific signal for AnxA1 in PMN ectosomes, and not PBMCs, was observed by FACS analysis. Western blot analysis confirmed the presence of a distinct double band at 33 and 37 kDa, likely corresponding to the cleaved and uncleaved forms of the protein. Mass spectrometry further confirmed the presence of AnxA1 as the protein with the highest MASCOT score upon in-gel digestion and analysis of the 37kDa band. Importantly, the anti-inflammatory effect of PMN-derived ectosomes was studied using a flow chamber apparatus, where a marked attenuation of cell rolling and adhesion was observed upon 5-10 min pre-incubation with 40-50 thousand ectosomes (>50% reduction; n=3, P<0.05). Of interest, this inhibitory effect was partially blocked by a neutralizing antibody against the AnxA1 receptor (also termed FPRL1) as well as by a peptide antagonist (peptide WRWWWW). These results indicate that AnxA1 is likely to be responsible for the inhibitory properties of PMN-derived ectosomes; we propose this represents a novel and exciting anti-inflammatory mechanism for the roles played by this endogenous anti-inflammatory mediator and may have potential clinical implications.