Fig. 5.

Ouabain effect on plasma membrane amounts of CFTR and NKCC1 in ADPKD monolayers. Confluent monolayers of ADPKD cells were treated without and with ouabain (3 nM). Following treatment, CFTR (a) and NKCC1 (b) levels at the plasma membrane were determined. Membrane proteins from ADPKD cell monolayers were isolated by biotinylation and streptavidin immunoprecipitation for CFTR or by isolating membrane fractions from the cells for NKCC1. Samples were analyzed by SDS-PAGE and immunoblot using antibodies specific for human CFTR and NKCC1, respectively. Bars represent membrane protein expression for each treatment relative to untreated control levels and normalized to total protein present in the sample, determined by Ponceau S. Values are expressed as the mean ± SEM of four determinations using cells from different ADPKD kidneys. Representative immunoblots are shown below the graph. *P < 0.05 versus untreated control