Quantitative Measurement of Urinary Podocalyxin by Custom-made Capture ELISA and Its Potential Application to Monitor Therapeutic Responses of Patients with Nephrotic Syndrome

Nephrotic syndrome, one of the most common manifestations of renal disorders encountered in daily nephrological clinics, is characterized with profound proteinuria. Proteinuria usually herald significant glomerular epithelial cell or podocyte injury. Podocyte presence in urine has been linked to some specific glomerular disease under several pathologic conditions. Podocalyxin (PC) is expressed at apical plasma membrane of podocyte and has been considered to contribute to the organization of foot processes as well as glomerular charge barrier through its extensive negative charge. We observed that through ELISA analysis urine sample from patients with diabetic nephropathy showed a relative higher PC level than that from healthy controls. We thus investigated if presence of the essential podocyte-associated membrane protein or PC itself in urine could represent the correlation with glomerular diseases or with patients’ prognosis. In this study we established a delicate capture ELISA kit specific to detect urinary PC. During the establishment we found to extract urinary PC required detergent contribution in ELISA assay. This might respond to Pisitkun et al. study that PC could reach urine through low-density membrane vesicles. After applying the custom-made capture ELISA kit to test urine samples from patients with clinically characterized nephrotic syndromes, including membranous nephropathy (MN), minimal change disease (MCD) and focal segmental glomerulosclerosis (FSGS), PC level was observed to be significantly lesser in MCD/FSGS group than in MN group at basal time (mean ± SEM: 2.0 ± 0.6, 5.7 ± 1.6, P< 0.05) before medical treatment. In addition PC level showed an increase at sixth month in MCD/FSGS group (2.0 ± 0.6 at 0 month, 4.3 ± 1.8 at 6 month, P< 0.1) during treatment course. In MN group this undulation of PC level was not observed. We explained the alteration of urinary PC level in MCD/FSGS group should account for the histological podocyte injury. In podocyte disease group, PC loss in the urine might be a prior step than the occurrence of foot processes impairment and its observation through biopsy. After patients received therapy, vigorous PC expression by podocyte might direct its sudden increase in urine due to the fact that plausible unstable and incomplete recovery of podocyte status yet.