Calcium phosphate deposits in domes of human pancreatic adenocarcinoma (Capan-1) cell cultures

Abstract

Human pancreatic cells of the Capan-1 line form domes in culture during the stationary growth stage. The domes are thought to be a result of the transport of water and electrolytes by the Capan-1 cells. In older Capan-1 cultures, the epithelial sheets formed thickenings from several layers of cells of which the outermost ones were joined by tight type junctions. In the intracellular space, deposits of insoluble calcium salts were observed. Culture of Capan-1 cells in the presence of fibroblasts prolonged survival of the cultures with intact domes for more than 80 days. The Capan-1 cells proliferated forming multilayers and closed cavities which we called super-domes. X-ray spectrometry and electron diffraction analysis showed that the abundant deposits inside these cavities consisted of calcium phosphate in an apatite structure. The number of these deposits increased with time in culture, and they appeared to be formed at the sites of contact with an extracellular matrix consisting of cell debris. Deposits were not observed within the culture medium. Cells from domes were stained cytochemically for ATPases and alkaline phosphatases and examined by light and electron microscopy. The Capan-1 cells surrounding the domes were differentiated, polarized cells containing placental type alkaline phosphatases on their apical membranes and Ca 2+-ATPases on their basolateral membranes. These enzymes were thought to play a role in the accumulation of phosphate and Ca 2+ ions in the dome cavities, which then formed crystals in the presence of organic compounds produced by lysis of cells of the deepest layers of the super-domes. The crystals of hydroxyapatite observed in standard Capan-1 cell cultures and those cocultured with fibroblasts were assumed to be a result of transepithelial transport of Ca 2+ and phosphate ions by these cells.