Supplementary Material for: Elevated Levels of Urinary Extracellular Vesicle Fibroblast-Specific Protein 1 in Patients with Active Crescentic Glomerulonephritis

<b><i>Background/Aims:</i></b> Extracellular vesicles (EVs), including exosomes, are present in various bodily fluids, including urine. We and others previously reported that cells expressing fibroblast-specific protein 1 (FSP1) accumulate within damaged glomeruli, and that urinary FSP1, as well as urinary soluble CD163, could potentially serve as a biomarker of ongoing glomerular injury. <b><i>Methods:</i></b> To test that idea, we collected urine samples from 37 patients with glomerular disease; purified the urinary EVs; characterized them using Nanosight, western blotting, and immunoelectron microscopy; and determined FSP1 and soluble CD163 levels using enzyme-linked immunosorbent assays. <b><i>Results:</i></b> Deemed to be mainly exosomes based on their size distribution, the EVs in urine contained FSP1, and a portion of the FSP1-positive vesicles was also positive for podocalyxin. FSP1 levels in urinary EVs were (1) positively correlated with rates of biopsy-proven cellular crescent formation (<i>r</i> = 0.562, <i>p</i> &lt; 0.001) and total crescent formation (<i>r</i> = 0.448, <i>p</i> = 0.005) among total glomeruli; (2) significantly higher in patients with cellular crescents affecting 20% or more of their glomeruli than in those with fewer affected glomeruli (<i>p</i> = 0.003); and (3) significantly decreased after glucocorticoid and immunosuppressant therapy (<i>p</i> &lt; 0.05). A positive correlation between FSP1 levels in urinary EVs and urinary soluble CD163 levels was confirmed (<i>r</i> = 0.367, <i>p</i> &lt; 0.05). <b><i>Conclusion:</i></b> These data suggest that a portion of urinary FSP1 is secreted as EVs originating from podocytes, and that FSP1 levels reflect active and ongoing glomerular injury and disease activity, such as cellular crescent formation.