Vectorial Bicarbonate Transport by Capan-1 Cells: a Model for Human Pancreatic Ductal Secretion

Abstract

Human pancreatic ducts secrete a bicarbonate-rich fluid but our knowledge of the secretory process is based mainly on studies of animal models. Our aim was to determine whether the HCO₃^- transport mechanisms in a human ductal cell line are similar to those previously identified in guinea-pig pancreatic ducts. Intracellular pH was measured by microfluorometry in Capan-1 cell monolayers grown on permeable filters and loaded with BCECF. Epithelial polarization was assessed by immunolocalization of occludin. Expression of mRNA for key electrolyte transporters and receptors was evaluated by RT-PCR. Capan-1 cells grown on permeable supports formed confluent, polarized monolayers with well developed tight junctions. The recovery of pH_i from an acid load, induced by a short NH₄⁺ pulse, was mediated by Na⁺-dependent transporters located exclusively at the basolateral membrane. One was independent of HCO₃^- and blocked by EIPA (probably NHE1) while the other was HCO₃^--dependent and blocked by H₂DIDS (probably pNBC1). Changes in pH_i following blockade of basolateral HCO₃^- accumulation confirmed that the cells achieve vectorial HCO₃^- secretion. Dose-dependent increases in HCO₃^- secretion were observed in response to stimulation of both secretin and VPAC receptors. ATP and UTP applied to the apical membrane stimulated HCO₃^- secretion but were inhibitory when applied to the basolateral membrane. HCO₃^- secretion in guinea-pig ducts and Capan-1 cell monolayers share many common features, suggesting that the latter is an excellent model for studies of human pancreatic HCO₃^- secretion.

Purinergic receptors in the endocrine and exocrine pancreas

“…In polarised monolayers, ATP and UTP applied luminally stimulated HCO À 3 secretion (measured as decrease in pH i ), while applied basolaterally they inhibited HCO À 3 secretion [104]. This is in agreement with studies on rat and guinea pig pancreatic ducts (see above).…”

“…This was in fact found in isolated guinea pig pancreatic ducts, where the basolateral application of UTP decreased secretion [103]. A similar effect of UTP on secretory processes was also detected in polarised Capan-1 cells, which are epithelial cells derived from pancreatic adenocarcinoma [104]. RT-PCR analysis of the single rat ducts revealed that they express Ca 2+ -activated K + channels of the intermediate conductance (IK also known as SLO, Maxi K and KCNMA1) and big conductance (BK also known as SK4, KCNN4) [105].…”

“…These findings implicate distinct roles of CD39 and purinergic signalling in tissue remodelling, fibrogenesis and perhaps tumour development. Future studies will need to clarify apparent lack and disagreement in immunolocalization of NTPDases [128,163] and P2 receptors in the parenchyma of the human pancreas and human duct cell lines as well as their molecular identity [104]. At least for P2 receptors, numerous functional studies on human and animal ducts show that they are present on the epithelium (see above).…”

Purinergic signalling in the pancreas in health and disease

“…As in other epithelia, P2 receptor localisation is difficult to reveal, as the same receptor type can be localised to both luminal and basolateral membranes, though coupled to different ion transporters (Novak 2008(Novak , 2011 (Ishiguro et al 1999, Steward et al 2005, Szücs et al 2006. P2X 7 receptors, most likely luminal, are cation channels but also decrease intracellular pH, possibly reflecting HCO K 3 secretion (Henriksen & Novak 2003).…”

Secretin

“…Secretin action has also been studied in monolayer cultures of canine and bovine pancreatic ducts (26,125) and in Capan-1 cells which are derived from a human tumor but which form a tight monolayer and possess secretin receptors (103,112). Only basic studies have been carried out to date but secretin will increase cAMP and ion transport in these monolayers.…”