Proteomic Profile of Urinary Extracellular Vesicles Identifies AGP1 as a Potential Biomarker of Primary Aldosteronism

Abstract

Context: Primary aldosteronism (PA) represents 6% to 10% of all essential hypertension patients and is diagnosed using the aldosterone-to-renin ratio (ARR) and confirmatory studies. The complexity of PA diagnosis encourages the identification of novel PA biomarkers. Urinary extracellular vesicles (uEVs) are a potential source of biomarkers, considering that their cargo reflects the content of the parent cell.

Objective: We aimed to evaluate the proteome of uEVs from PA patients and identify potential biomarker candidates for PA.

Methods: Second morning spot urine was collected from healthy controls (n = 8) and PA patients (n = 7). The uEVs were isolated by ultracentrifugation and characterized. Proteomic analysis on uEVs was performed using LC-MS Orbitrap.

Results: Isolated uEVs carried extracellular vesicle markers, showed a round shape and sizes between 50 and 150 nm. The concentration of uEVs showed a direct correlation with urinary creatinine (r = 0.6357; P = 0.0128). The uEV size mean (167 ± 6 vs 183 ± 4nm) and mode (137 ± 7 vs 171 ± 11nm) was significantly smaller in PA patients than in control subjects, but similar in concentration. Proteomic analysis of uEVs from PA patients identified an upregulation of alpha-1-acid glycoprotein 1 (AGP1) in PA uEVs, which was confirmed using immunoblot. A receiver operating characteristic curve analysis showed an area under the curve of 0.92 (0.82 to 1; P = 0.0055).

Conclusion: Proteomic and further immunoblot analyses of uEVs highlights AGP1 as potential biomarker for PA.

Keywords: aldosterone-to-renin ratio; hypertension; proteomics; urine; vesicles.