title: Identification and biochemical localization of a Na-K-Cl cotransporter in the human placental cell line BeWo.
abstract: Several transport systems mediating the placental transport of Na, K and Cl have been described, but whether the trophoblast membrane also expresses a Na-K-Cl cotransporter that mediates the coupled movement of all three ions remains unclear. Here we show that BeWo cells, a human trophoblastic cell line, exhibit bumetanide-sensitive (86)Rb (a K surrogate) uptake. Entry via this route accounts for approximately 17% of the (86)Rb influx with the remainder being mediated largely via the Na,K-ATPase. The activity of the bumetanide-sensitive transporter was rapidly elevated (>40%) upon subjecting cells to an acute hyperosmotic challenge signifying a potential role in cell volume regulation. Antibodies to the Na-K-Cl cotransporter identified a single band of approximately 200 kDa on Western blots of fractionated BeWo membranes. This immunoreactivity colocalized with that of the Na,K-ATPase (a basal membrane marker), but was absent from membranes enriched with placental alkaline phosphatase (an apical membrane marker). These findings show for the first time, that a Na-K-Cl cotransporter is expressed in a human placental cell line which may be involved in regulating trophoblast cell volume.
authors.affiliation: Department of Anatomy and Physiology, Medical Sciences Institute/Wellcome Trust Biocentre Complex, Dundee, DD1 4HN, Scotland.
authors.forename: H
authors.initials: H
authors.lastname: Zhao
authors.forename: H S
authors.initials: HS
authors.lastname: Hundal
mesh_terms: D015153:Blotting, Western; D001921:Brain / Q000378:metabolism; D002034:Bumetanide / Q000493:pharmacokinetics; D002352:Carrier Proteins / Q000096:biosynthesis* / Q000737:chemistry*; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004232:Diuretics / Q000493:pharmacokinetics; D004305:Dose-Response Relationship, Drug; D004591:Electrophoresis, Polyacrylamide Gel; D004791:Enzyme Inhibitors / Q000493:pharmacokinetics; D006801:Humans; D008564:Membrane Potentials; D008861:Microsomes / Q000378:metabolism; D009995:Osmosis; D010042:Ouabain / Q000493:pharmacokinetics; D010920:Placenta / Q000378:metabolism* / Q000502:physiology; D011188:Potassium / Q000493:pharmacokinetics*; D020033:Protein Isoforms; D012413:Rubidium / Q000493:pharmacokinetics; D028021:Sodium-Potassium-Chloride Symporters; D000254:Sodium-Potassium-Exchanging ATPase / Q000737:chemistry; D013347:Subcellular Fractions; D013997:Time Factors
chemical_list: D002352:Carrier Proteins; D004232:Diuretics; D004791:Enzyme Inhibitors; D020033:Protein Isoforms; D028021:Sodium-Potassium-Chloride Symporters; D002034:Bumetanide; D010042:Ouabain; D000254:Sodium-Potassium-Exchanging ATPase; D012413:Rubidium; D011188:Potassium