Immortalization of bovine pancreatic duct epithelial cells.
Pancreatic duct cell lines have been isolated from a number of animal and human tumors, but none appear to express ion transport properties expected for differentiated pancreatic duct epithelial cells. We sought to generate an immortalized ductal cell line from well-differentiated primary cultures of bovine pancreatic duct epithelium. Epithelial cells from the main duct of the bovine pancreas were isolated and immortalized by transfection with a DNA construct encoding simian virus 40 large T antigen. A single clone (BPD1) survived negative selection and was maintained in culture for > 100 passages over 2 yr. The cells grow readily in culture as monolayers and express several properties characteristic of differentiated pancreatic ductal epithelium. The cells do not appear to form a functional tight junction complex, since the transepithelial resistance of the monolayer cultures grown on a permeable support is < 10 omega.cm2. Northern blot analysis revealed that the cells continue to express simian virus 40 large T antigen and contain significant levels of mRNA for proteins thought to be important in transepithelial bicarbonate secretion [carbonic anhydrase II, Cl-/HCO3- exchanger, Na+/H+ exchanger, and cystic fibrosis transmembrane conductance regulator (CFTR)]. In vivo pancreatic ductal secretion is stimulated by the peptide hormone secretin. The secretin receptor is expressed and functionally coupled to adenylate cyclase in the immortalized cells, since secretin caused a dose-dependent accumulation of adenosine 3'5'-cyclic monophosphate (cAMP; approximately 20-fold increase over basal levels) with a mean effective concentration of 15 nM. Elevation of intracellular cAMP by exposure of the cells to forskolin (10 microM) or secretin (0.1 microM) increase plasma membrane Cl- permeability, most likely mediated by activation of CFTR. The results of these studies demonstrate that the pancreatic duct cell line (BPD1) retains several properties exhibited by the secretory epithelial cells that line the pancreatic ductal tree. This cell line should prove useful for studies of expression, function, and regulation of pancreatic duct cell proteins.
Marino
L R
LR
Department of Pediatrics, Case Western Reserve University, Cleveland, Ohio 44106, USA.
Cotton
C U
CU
D000818:Animals; D000952:Antigens, Polyomavirus Transforming / Q000235:genetics / Q000378:metabolism; D017920:Antiporters / Q000235:genetics; D002256:Carbonic Anhydrases / Q000235:genetics; D002417:Cattle; D002455:Cell Division; D002461:Cell Line, Transformed; D002462:Cell Membrane / Q000378:metabolism; D002463:Cell Membrane Permeability; D027963:Chloride-Bicarbonate Antiporters; D002999:Clone Cells; D019005:Cystic Fibrosis Transmembrane Conductance Regulator / Q000235:genetics; D004847:Epithelial Cells; D004848:Epithelium / Q000502:physiology; D015870:Gene Expression; D007477:Ions; D010183:Pancreatic Ducts / Q000166:cytology* / Q000502:physiology; D015398:Signal Transduction; D017923:Sodium-Hydrogen Exchangers / Q000235:genetics
D000952:Antigens, Polyomavirus Transforming; D017920:Antiporters; D027963:Chloride-Bicarbonate Antiporters; D007477:Ions; D017923:Sodium-Hydrogen Exchangers; D019005:Cystic Fibrosis Transmembrane Conductance Regulator; D002256:Carbonic Anhydrases