Complement receptor CD35 on single urinary extracellular vesicle as novel biomarker of sepsis-associated acute kidney injury

Abstract Sepsis-associated acute kidney injury (SA-AKI) portends severe health burden due to significant morbidity and mortality, while early diagnosis remains challenging. In this study, proximity-dependent barcoding assay (PBA) was established to profile the surface proteome of single urinary extracellular vesicle (uEV). Principle uEV clusters with unique function and origination were profiled in SA-AKI. Reduction of complement receptor CD35 on single uEV (CD35-uEV) was revealed as a novel biomarker from one of the main EV clusters with significant proportional differences. CD35-uEV demonstrated high diagnostic accuracy for SA-AKI (receiver operating characteristic-area under the curve (ROC-AUC), 0.98 in screening cohort (n=16), and 0.89 in validation cohort (n=134)). Besides, CD35-uEV enables identification of subclinical AKI (ROC-AUC, 0.84 in prospective cohort (n=72)) which was independent of other clinical parameters as validated by multivariate analysis (p<0.001). Moreover, CD35-uEV correlated closely with AKI severity which also predicts persistent AKI (ROC-AUC, 0.77), progression to AKD (ROC-AUC, 0.66), and mortality risks (ROC-AUC, 0.70). Integrative single-cell and spatial transcriptomics analysis identified that CD35-uEV originated from injured podocyte characterized with diminished CD35 expression. The combination of CD35-uEV with tubular injury biomarkers (TIMP2*IGFBP7) showed improved accuracy in identifying subclinical SA-AKI and prediction of severe stages. Overall, this study identified a novel biomarker on single uEV related to injured podocyte for early diagnosis and risk stratification of SA-AKI.