Urine exosomes from healthy and hypertensive pregnancies display elevated level of α-subunit and cleaved α- and γ-subunits of the epithelial sodium channel—ENaC

• Published in: Pflügers Archiv 2017-09, Vol.469 (9), p.1107-1119
• 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: Article
• Language: English
• Subjects: Adult; Aldosterone - urine; Biomedical and Life Sciences; Biomedicine; Cell Biology; Epithelial Sodium Channels - metabolism; Epithelial Sodium Channels - urine; Exosomes - metabolism; Female; Fibrinolysin - urine; Human Physiology; Humans; Hypertension - metabolism; Hypertension - urine; Kidney - metabolism; Molecular and Cellular Mechanisms of Disease; Molecular Medicine; Neurosciences; Potassium - urine; Pre-Eclampsia - metabolism; Pre-Eclampsia - urine; Pregnancy; Protein Subunits - metabolism; Protein Subunits - urine; Proteinuria - metabolism; Proteinuria - urine; Receptors; Sodium - urine; Urine - physiology
• 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), α-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 α-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 α-ENaC subunit while γ-subunit appeared predominantly in cleaved form independently of conditions and with a significant contribution from post-renal cleavage.