Treatment with an EV secretion inhibitor decreased the amount of CD63 and HA deposited, substantiating the role of EVs in nucleating mineral.15,38–40 We also observed substantial retention of Ca in the minimally mineralized early ECM, which is consistent with in vivo studies in children with CKD where Ca accumulation in the vessel wall precedes the development of overt calcification and where EVs are the main nidus for mineralization.37 This evidence concerns the gene CD63 and chronic kidney disease.