Urinary exosomes have been proposed as potential diagnostic tools. TNF 
superfamily cytokines and receptors may be present in exosomes and are expressed 
by proximal tubular cells. We have now studied the expression of selected TNF 
superfamily proteins in exosome-like vesicles from cultured human proximal 
tubular cells and human urine and have identified additional proteins in these 
vesicles by LC-MS/MS proteomics. Human proximal tubular cells constitutively 
released exosome-like vesicles that did not contain the TNF superfamily 
cytokines TRAIL or TWEAK. However, exosome-like vesicles contained 
osteoprotegerin (OPG), a TNF receptor superfamily protein, as assessed by 
Western blot, ELISA or selected reaction monitoring by nLC-(QQQ)MS/MS. 
Twenty-one additional proteins were identified in tubular cell exosome-like 
vesicles, including one (vitamin D binding protein) that had not been previously 
reported in exosome-like vesicles. Twelve were extracellular matrix proteins, 
including the basement membrane proteins type IV collagen, nidogen-1, agrin and 
fibulin-1. Urine from chronic kidney disease patients contained a higher amount 
of exosomal protein and exosomal OPG than urine from healthy volunteers. 
Specifically OPG was increased in autosomal dominant polycystic kidney disease 
urinary exosome-like vesicles and expressed by cystic epithelium in vivo. In 
conclusion, OPG is present in exosome-like vesicles secreted by proximal tubular 
epithelial cells and isolated from Chronic Kidney Disease urine.