Urine exosomes from healthy and hypertensive pregnancies display elevated level of [alpha]-subunit and cleaved [alpha]- and [gamma]-subunits of the epithelial sodium channel--ENaC

• Published in: Pflügers Archiv Vol. 469; no. 9; p. 1107
• Main Authors: Nielsen, Maria R, Frederiksen-møller, Britta, Zachar, Rikke, Jørgensen, Jan S, Hansen, Mie R, Ydegaard, Rikke, Svenningsen, Per, Buhl, Kristian, Jensen, Boye L
• Format: Journal Article
• Language: English
• Published: Heidelberg Springer Nature B.V 01-09-2017
• Subjects: Aldosterone; Angiotensin; Catheters; Epitopes; Excretion; Exosomes; Furin; Immunoblotting; Kidneys; Pelvis; Plasmin; Pre-eclampsia; Preeclampsia; Pregnancy; Proteinuria; Reabsorption; Renal cortex; Renin; Sodium; Ultracentrifugation; Urine
• ISSN: 0031-6768; 1432-2013
• DOI: 10.1007/s00424-017-1977-z

Preeclampsia is characterized by hypertension, proteinuria, suppression of plasma renin-angiotensin-aldosterone, and impaired urine sodium excretion. Aberrantly filtered plasmin in urine may activate proteolytically the γ-subunit of the epithelial sodium channel (ENaC) and promote Na+ reabsorption and urine K+ loss. Plasma and urine was sampled from patients with preeclampsia, healthy pregnant controls and non-pregnant women, and from patients with nephrostomy catheters. Aldosterone concentration, urine plasminogen, and protein were determined. Exosomes were isolated by ultracentrifugation. Immunoblotting was used to detect exosome markers; γ-ENaC (two different epitopes within the inhibitory peptide tract), [alpha]-ENaC, and renal outer medullary K-channel (ROMK) and compared with human kidney cortex homogenate. Urine total plasmin(ogen) was significantly increased in preeclampsia, plasma and urine aldosterone was higher in pregnancy compared to non-pregnancy, and the urine Na/K ratio was lower in preeclampsia compared to healthy pregnancy. Exosome markers ALIX and AQP-2 were stably associated with exosomes across groups. Exosomal [alpha]-ENaC-subunit migrated at 75 kDa and dominantly at 50 kDa and was significantly elevated in pregnancy. In human kidney cortex tissue and two of four pelvis catheter urine, ~90-100 kDa full-length γ-ENaC was detected while no full-length γ-ENaC but 75, 60, and 37 kDa variants dominated in voided urine exosomes. There was no difference in γ-ENaC protein abundances between healthy pregnancy and preeclampsia. ROMK was detected inconsistently in urine exosomes. Pregnancy and preeclampsia were associated with increased abundance of furin-cleaved [alpha]-ENaC subunit while γ-subunit appeared predominantly in cleaved form independently of conditions and with a significant contribution from post-renal cleavage.