DOG PANCREATIC DUCT EPITHELIAL-CELLS - LONG-TERM CULTURE AND CHARACTERIZATION

Epithelial cells, isolated from a normal dog pancreatic duct, were gro wn on collagen-coated culture inserts suspended above a feeder layer o f myofibroblasts. The cells were examined by transmission electron mic roscopy, immunohistochemistry, cytogenetics, and flow cytometry. In ad dition, the constitutive and agonist-stimulated mucin secretion of the se cells was studied using a [H-3]N-acetyl-D-glucosamine labeling assa y, and the stimulation of intracellular cAMP was measured Cells grown on inserts with a feeder layer developed into confluent monolayers con sisting of strictly polarized columnar epithelial cells with prominent microvilli, intercellular junctions, and normal chromosomal character istics. They could be passaged repeatedly without a detectable alterat ion in their morphology. The cells could also be grown on organotypic cultures, resulting in further differentiated cells simulating in vivo morphology. Immunohistochemistry demonstrated the presence of carboni c anhydrase II in these cells. Cells treated with vasoactive intestina l peptide, epinephrine, and dibutyryl-cAMP demonstrated a marked incre ase in mucin secretion compared with controls. In parallel experiments , MP and epinephrine significantly, increased intracellular cAMP. In c onclusion we have developed a pancreatic epithelial cell preparation w ith morphology, cytokinetics, chromosomal, and DNA analyses characteri stic of normal cells. Similar to normal columnar epithelial cells, the se pancreatic duct cells secreted mucin constitutively and responded t o agonist by increasing secretion via a cAMP-mediated pathway. They al so contained carbonic anhydrase, which indicates that the cells are ca pable of secreting bicarbonate.