Vascular calcification (VC) is a strong predictor of cardiovascular risk, particularly in chronic kidney disease (CKD) patients, associated with increased vascular stiffness, pulse pressure, left ventricular hypertrophy and atherosclerotic plaque burden [1]. VC is currently accepted as a highly controlled multifactorial process, where the release of extracellular vesicles (EVs) with calcification capacity plays an essential role in mediating cell-induced matrix mineralization [2]. Still, questions considering mechanisms of EVs deposition in the extracellular space, and its relationship with vesicle origin, loading, calcifying capacity and role in intercellular communication remain elusive. In this work we established a proteomic approach to characterize extracellular matrix (ECM)-deposited and cell media (CM) released EVs, from an in vitro model of VC consisting of primary vascular smooth muscle cells (VSMCs), bringing new knowledge into their specific characteristics and cargo.