Urinary Extracellular Vesicles as a Source of Biomarkers for Kidney Allograft Rejection: A Systematic Review

Abstract

BACKGROUND: Kidney transplant rejection remains a leading cause of graft loss, and diagnosis still depends largely on invasive biopsies. Urinary extracellular vesicles (uEVs) offer a potential noninvasive alternative for detecting allograft rejection through molecular profiling of vesicle cargo. METHODS: We conducted a systematic search of major databases for studies published up to November 2025 evaluating uEV biomarkers for kidney transplant rejection. We extracted data on analytical methods and diagnostic performance and assessed clinical applicability. Quality assessment was performed using QUADAS-2. RESULTS: Sixteen studies met the inclusion criteria. Studies employed diverse uEV isolation methods: ultracentrifugation (n = 7), commercial kits (n = 7), size exclusion chromatography (n = 1), and direct-analysis flow cytometry-based approach (n = 1). Analytical platforms included proteomics (n = 9), transcriptomics (n = 6), and genomics (n = 1). Only 1 study examined pediatric recipients. QUADAS-2 assessment revealed generally low risk of bias, though concerns regarding blinding and threshold prespecification were noted in 13 studies. Identified biomarkers predominantly reflected immune activation pathways, including interferon/JAK-STAT signaling, chemokine/CXCR3 axis, antigen presentation, TLR/inflammasome activation, and complement cascade components. Several studies achieved an area under the curve >0.75 in externally validated cohorts, with high negative predictive values (>90%) and positive predictive values of 38%-63%. Key limitations included small sample sizes, cross-sectional designs, nonstandardized protocols, and a lack of normalization. uEVs harbor molecular signatures that could complement current diagnostic approaches for kidney transplant rejection. Biomarkers predominantly reflect coordinated immune activation integrated with metabolic and tissue-specific adaptations. However, clinical translation requires standardized isolation protocols, multicenter validation studies, and expanded pediatric research. TRIAL REGISTRATION: PROSPERO 2025 CRD420251138942. https://www.crd.york.ac.uk/PROSPERO/view/CRD420251138942.