title: Glibenclamide stimulates fluid secretion in rodent cholangiocytes through a cystic fibrosis transmembrane conductance regulator-independent mechanism.
abstract: OBJECTIVE
  Progressive liver disease is a severe complication of cystic fibrosis, a genetic disease characterized by impaired epithelial adenosine 3',5'-cyclic monophosphate-dependent secretion caused by mutations in the cystic fibrosis transmembrane conductance regulator (CFTR). In the liver, CFTR is expressed in cholangiocytes and regulates the fluid and electrolyte content of the bile. Glibenclamide, a sulfonylurea and a known CFTR inhibitor, paradoxically stimulates cholangiocyte secretion. We studied the molecular mechanisms underlying this effect and whether glibenclamide could restore cholangiocyte secretion in cystic fibrosis.
  
  
  METHODS
  NRC-1 cells, freshly isolated rat cholangiocytes, isolated rat biliary ducts, and isolated biliary ducts from CFTR-defective mice (Cftr tm1Unc ) were used to study fluid secretion (by video-optical planimetry), glibenclamide-induced secretion (by high-performance liquid chromatography in cell culture medium), intracellular pH and intracellular Ca 2+ concentration transients [2'7'-bis(2-carboxyethyl)-5,6,carboxyfluorescein-acetoxymethylester and Fura-2 f-AM (5-Oxazolecarboxylic acid, 2-(6-(bis(2-((acetyloxy)methoxy)-2-oxoethyl)amino)-5-(2-(2-(bis(2-((acetyloxy)methoxy)-2-oxoethyl)amino)-5-methylphenoxy)ethoxy)-2-benzofuranyl)-, (acetyloxy)methyl ester) microfluorometry], gene expression (by reverse-transcription polymerase chain reaction), and changes in membrane capacitance (by patch-clamp experiments).
  
  
  RESULTS
  Stimulation of cholangiocyte secretion by glibenclamide and tolbutamide required Cl - and was mediated by the sulfonylurea receptor 2B. Glibenclamide-induced secretion was blocked by inhibitors of exocytosis (colchicine, wortmannin, LY294002, and N -ethylmaleimide) and by inhibitors of secretory granule acidification (vanadate, bafilomycin A1, and niflumic acid) but was Ca 2+ and depolarization independent; membrane capacitance measurements were consistent with stimulation of vesicular transport and fusion. Glibenclamide, unlike secretin and forskolin, was able to stimulate secretion in Cftr tm1Unc mice, thus indicating that this secretory mechanism was preserved.
  
  
  CONCLUSIONS
  The ability of glibenclamide to stimulate secretion in CFTR-defective mice makes sulfonylureas a model class of compounds to design drugs useful in the treatment of cystic fibrosis with liver impairment and possibly of other cholestatic diseases.
authors: Department of Meidcal and Surgical Sciences, University of Padova, Italy.
authors: Carlo
authors: C
authors: Spirlì
authors: Romina
authors: R
authors: Fiorotto
authors: Lei
authors: L
authors: Song
authors: Joseph
authors: J
authors: Santos-Sacchi
authors: Lajos
authors: L
authors: Okolicsanyi
authors: Sara
authors: S
authors: Masier
authors: Loretta
authors: L
authors: Rocchi
authors: Maria Pia
authors: MP
authors: Vairetti
authors: Marina
authors: M
authors: De Bernard
authors: Saida
authors: S
authors: Melero
authors: Tullio
authors: T
authors: Pozzan
authors: Mario
authors: M
authors: Strazzabosco
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000378:metabolism; D000818:Animals; D001652:Bile Ducts / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D001826:Body Fluids / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002460:Cell Line; D003550:Cystic Fibrosis / Q000188:drug therapy* / Q000378:metabolism / Q000503:physiopathology; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000378:metabolism*; D025802:Electric Capacitance; D004553:Electric Conductivity; D005905:Glyburide / Q000494:pharmacology*; D007004:Hypoglycemic Agents / Q000494:pharmacology*; D008099:Liver / Q000187:drug effects / Q000502:physiology; D051379:Mice; D018181:Mice, Inbred CFTR; D018408:Patch-Clamp Techniques; D015221:Potassium Channels / Q000378:metabolism; D024661:Potassium Channels, Inwardly Rectifying / Q000378:metabolism; D051381:Rats; D011955:Receptors, Drug / Q000378:metabolism; D064233:Sulfonylurea Receptors
chemical_list: D018528:ATP-Binding Cassette Transporters; D007004:Hypoglycemic Agents; D015221:Potassium Channels; D024661:Potassium Channels, Inwardly Rectifying; D011955:Receptors, Drug; D064233:Sulfonylurea Receptors; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D005905:Glyburide; D002118:Calcium