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OBJECTIVES: Increased activity of the epithelial sodium channel (ENaC) in the kidneys may explain the coupling between proteinuria, edema, suppressed aldosterone and hypertension in preeclampsia. Preeclamptic women excrete plasminogen-plasmin in urine. In vitro, plasmin increases the activity of ENaC by proteolytic cleavage of the γ-subunit ectodomain and release of a 43-aminoacid inhibitory tract from the channel. Exosomes are membrane vesicles released into the urine from apical membranes of the kidney epithelial cells.OBJECTIVES: (1) To investigate if the proteolytic state of the ENaC γ-subnit can be studied in urine exosomes from pregnant women. (2) To investigate if the ENaC γ-subunit ectodomain is abnormally activated by proteolysis in preeclamptic women.METHODS: 100 mL spot urine samples from 14 preeclamptic women, 17 pregnant women and 9 non-pregnant women were collected with protease inhibitors. Plasmin/plasminogen was measured. Exosomes were recovered by ultracentrifugation at 220,000×g at 4°C for 100 min. The exosome fraction was used for western blotting with a newly developed monoclonal antibody, mAb3C7, directed against the "inhibitory" tract in γ-ENaC. Aquaporin-2 (AQP2) was used as a positive control for the presence of collecting duct membrane.RESULTS: Urine plasmin-plasminogen/creatinine ratio was increased in the preeclampsia group (pCONCLUSIONS: It is possible to examine collecting duct transport proteins in urine exosome from pregnant women including γ-ENaC, 2) Urine exosome fraction displays a variable pattern of γ-ENaC signal with a predominance of cleaved forms in both normal and preeclamptic women.DISCLOSURES: M.R. Nielsen: None. M. Rytz: None. B. Frederiksen-Møller: None. P. Svenningsen: None. R.M. Zachar: None. J.S. Jørgensen: None. B.L. Jensen: None.