Plasmin in Nephrotic Urine Activates the Epithelial Sodium Channel

TLDR

Proteinuria and increased renal reabsorption of NaCl characterize the nephrotic syndrome. We identified plasmin as a urinary serine protease by matrix‑assisted laser desorption/ionization time‑of‑flight mass spectrometry. Protein‑rich urine from nephrotic rats and patients activates ENaC in cultured cells and oocytes via urinary serine protease activity, with purified plasmin directly stimulating ENaC currents and urokinase‑mediated conversion of plasminogen to plasmin enhancing sodium transport through cleavage of an inhibitory peptide from the ENaC γ subunit, indicating that a defective filtration barrier permits proteolytic enzymes to activate ENaC.

Abstract

Proteinuria and increased renal reabsorption of NaCl characterize the nephrotic syndrome. Here, we show that protein-rich urine from nephrotic rats and from patients with nephrotic syndrome activate the epithelial sodium channel (ENaC) in cultured M-1 mouse collecting duct cells and in <i>Xenopus laevis</i> oocytes heterologously expressing ENaC. The activation depended on urinary serine protease activity. We identified plasmin as a urinary serine protease by matrix-assisted laser desorption/ionization time of-flight mass spectrometry. Purified plasmin activated ENaC currents, and inhibitors of plasmin abolished urinary protease activity and the ability to activate ENaC. In nephrotic syndrome, tubular urokinase-type plasminogen activator likely converts filtered plasminogen to plasmin. Consistent with this, the combined application of urokinase-type plasminogen activator and plasminogen stimulated amiloride-sensitive transepithelial sodium transport in M-1 cells and increased amiloride-sensitive whole-cell currents in <i>Xenopus laevis</i> oocytes heterologously expressing ENaC. Activation of ENaC by plasmin involved cleavage and release of an inhibitory peptide from the ENaC γ subunit ectodomain. These data suggest that a defective glomerular filtration barrier allows passage of proteolytic enzymes that have the ability to activate ENaC.