Albuminuria in kidney transplant recipients is associated with increased urinary serine proteases and activation of the epithelial sodium channel

Abstract

Albuminuria predicts adverse renal outcome in kidney transplant recipients. The present study addressed the hypothesis that albuminuria is associated with increased urine serine proteases with the ability to activate the epithelial sodium channel (ENaC) and with greater extracellular volume and higher blood pressure. In a cross-sectional design, kidney transplant recipients with ( n = 18) and without ( n = 19) albuminuria were included for office blood pressure measurements, estimation of volume status by bioimpedance, and collection of spot urine and plasma samples. Urine was analyzed for serine proteases and for the ability to activate ENaC current in vitro. Urine exosome protein was immunoblotted for prostasin and γ-ENaC protein. In the present study, it was found that, compared with nonalbuminuria (8.8 mg/g creatinine), albuminuric (1,722 mg/g creatinine) kidney transplant recipients had a higher systolic and diastolic blood pressure, despite receiving significantly more antihypertensives, and a greater urinary total plasminogen, active plasmin, active urokinase-type plasminogen activator, and prostasin protein abundance, which correlated significantly with u-albumin. Fluid overload correlated with systolic blood pressure, urinary albumin/creatinine, and plasminogen/creatinine. Urine from albuminuric kidney transplant recipients evoked a greater amiloride- and aprotinin-sensitive inward current in single collecting duct cells (murine cell line M1). γENaC subunits at 50 and 75 kDa showed increased abundance in urine exosomes from albuminuric kidney transplant recipients when compared with controls. These findings show that albuminuria in kidney transplant recipients is associated with hypertension, ability of urine to proteolytically activate ENaC current, and increased abundance of γENaC. ENaC activity could contribute to hypertension and adverse outcome in posttransplant proteinuria.

Roles for Exosome in Various Kidney Diseases and Disorders

“…Additionally, the information obtained from exosomes also leads to better understanding of the mechanisms and pathogenic events after kidney transplantation. For example, the increased levels of 50-kDa and 75-kDa subunits of g-ENaC were found in urinary exosomes derived from albuminuric kidney transplant recipients and was associated with hypertension (Hinrichs et al, 2018). Moreover, the decreased level of AQP-2 was detected in urinary extracellular vesicles (including exosomes) 1-day after kidney transplantation together with increased urinary volume and decreased urinary osmolality (Oshikawa-Hori et al, 2019).…”

Dietary Na+intake in healthy humans changes the urine extracellular vesicle prostasin abundance while the vesicle excretion rate, NCC, and ENaC are not altered

“…ENaC is activated by aldosterone by complex mechanisms involving apical targeting, increased expression, suppressed ubiquitination (9,(21)(22)(23)(24)(25) and proteolytical activation (8,11,14,20,26,27,29,31,3). ENaC has been detected in uEVs by mass spectrometry (17,32) and immunoblotting (2,18,42) and its urinary excretion is increased upon low dietary Na + intake or administration of aldosterone in humans (32). Proteolytic activation is believed to contribute to both physiological and pathological regulation of ENaC and is well established in experimental animals (37,39).…”

Urinary Plasmin(ogen): New Predictor of Hypertension?

“…Furthermore, urine from patients with diabetic (types 1 and 2) nephropathy or proteinuric renal transplant recipients showed increased ENaC activity in cultured collecting duct cells and contained elevated cleaved ENaC-γ. 2 , 3 , 5 Finally, administration of the serine protease inhibitor aprotinin normalized urinary serine protease activity and cleaved ENaC-γ expression and sodium excretion in experimental nephrotic syndrome. 8 Taken together, these findings suggest that elevated tubule fluid plasminogen/urokinase/plasmin contributes to sodium retention and hypertension in proteinuric kidney diseases.…”

“… 1 When the glomerular filtration barrier is compromised, plasminogen can reach the tubule lumen where, after proteolysis by urokinase, which is normally present in urine, plasmin is generated with resultant potential activation of ENaC. 1 Plasmin-mediated ENaC activation may be of pathological importance in that an increasing number of reports describe elevated urinary plasmin(ogen) (assays typically do not distinguish between plasminogen and plasmin) in patients with proteinuria and increased blood pressure, including type 2 diabetes mellitus with resistant hypertension, 2 type 1 diabetes, 3 hydronephrosis, 4 kidney transplants, 5 chronic kidney disease with overhydration, 6 preeclampsia, 7 and other conditions. Furthermore, urine from patients with diabetic (types 1 and 2) nephropathy or proteinuric renal transplant recipients showed increased ENaC activity in cultured collecting duct cells and contained elevated cleaved ENaC-γ.…”

“…Indeed, in this and several other studies, urine plasmin(ogen) directly correlated with urine albumin. 2 , 3 , 4 , 5 , 6 , 7 Furthermore, albuminuria is widely accepted to prognosticate adverse renal and cardiovascular outcomes; tubular luminal albumin may exert direct pathogenic actions and/or is associated with other pathogenic factors when the glomerular filtration barrier is compromised. Notably, uPl/Cr did not improve the prediction of primary or secondary outcomes in the current study beyond that of uAlb/Cr; in the one other uPl/Cr prognostication study, albuminuria was a stronger predictor of preeclampsia than uPl/Cr.…”