Abstract: SA-PO1014
Urinary Exosomes Quantitation of Renal Transporter Abundance Correlates with Function
Session Information
- Fluid and Electrolytes: Basic - II
October 27, 2018 | Location: Exhibit Hall, San Diego Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Fluid and Electrolytes
- 901 Fluid and Electrolytes: Basic
Authors
- Menezes, Cameron J., University of Chicago, Chicago, Illinois, United States
- Bergsland, Kristin J., University of Chicago, Chicago, Illinois, United States
- Worcester, Elaine M., University of Chicago, Chicago, Illinois, United States
- Coe, Fredric L., University of Chicago, Chicago, Illinois, United States
- Ko, Benjamin S., University of Chicago, Chicago, Illinois, United States
Background
Urinary exosomes have the potential to provide considerable insight into renal tubular function. However, estimates of urinary exosomes been qualitative or semi-qualitative using Western blotting. Therefore, we sought develop approaches to measure several relevant sodium and calcium transporters simultaneously in the urine from humans and then correlate them to function in humans.
Methods
4 healthy males were studied in a CRC setting. Fasting and fed urine and blood samples were collected hourly. Serum ultrafiltrate and urine calcium, magnesium, sodium, potassium, creatinine, and urine volume were measured. Lithium was measured in deproteinated serum and urine by atomic emission spectroscopy. Urinary exosomes were isolated via ultracentrifugation. NHE3, SLC26A6, NKCC2, NCC, TRPV5 and prostasin abundance were measured via ELISA at each collection point in triplicate and these values were compared with metabolic parameters of tubule function at each time point.
Results
The presence of enriched amounts of ALIX, CD63, and TSG-101 protein in the exosomal fraction compared to the various wash fractions was verified via Western blotting. NHE3, SLC26A6, NKCC2, NCC, TRPV5 and prostasin abundance were measured. NHE3 and fractional excretion of lithium are significantly correlated, indicating that proximal tubule function is related to the exosomal abundance of NHE3 (-0.66, p=0.001). Fully corrected for multiple comparisons, significant inverse correlations between the fractional excretion of distally delievered Ca and TRPV5 (-0.76, p=0.001) and FEDNa and prostasin (-0.78, p=0.001) were demonstated.
Conclusion
Our findings represent reliable and precise methods for quantitation of urinary exosomal abundance of the transport proteins NHE3, SLC26A6, NKCC2, NCC, TRPV5, and prostasin. The quantitation of urinary exosome renal transporter protein abundance allows for greater insight into human renal transport, allowing for direct measurements of markers of molecular solute transport. In this way, this system has the potential to further our understanding of molecular electrolyte physiology and disease in humans by confirming previous theories and finding new connections.
Funding
- NIDDK Support