Surface charge alterations in urinary extracellular vesicles mirror diabetic kidney disease pathophysiology

Urinary extracellular vesicles (uEVs) are an attractive source of biomarkers as their cargo of proteins, nucleic acids and lipids reflects cell pathophysiology. The physical properties of uEVs such as their electrical nature measured by the zeta potential (charge at the interface of vesicle and liquid) are seldom explored although their understanding may lead to translational applications of EVs in clinical settings. In diabetes and diabetic kidney disease (DKD), post‐translational modifications resulting from loss of glycocalyx (diabetes) and gain of extracellular matrix components (tissue fibrosis) could potentially affect the electrical charge of pathologically relevant EVs. DKD cohorts obtained by adhering to industry‐standard reproducible criteria were stratified into stable and progressive disease based on estimated glomerular filtration rate (eGFR) loss and compared to nonkidney disease diabetic and healthy controls. Surface charge analysis revealed a significant difference in the zeta potential of large uEVs between the diabetic and healthy cohorts, as well as between the diabetic and DKD‐progressive cohorts. These differences are mechanistically consistent with pathological alterations observed in heparan sulfate (HS) content. Accordingly, uEVs from DKD patients exhibited higher HS levels than those from diabetic individuals, as assessed by SP‐IRIS/ ExoView using HS‐binding capture. Comparative proteomic analysis further demonstrated an increased abundance of the sulfated glycosaminoglycan lumican in DKD uEVs relative to diabetes, consistent with its reported upregulation in early fibrotic stages of DKD. Notably, our findings indicate for the first time that the zeta potential of large uEVs may be predictive of the progression of diabetes into DKD.