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Characterization of H+ and HCO3- transporters in CFPAC-1 human pancreatic duct cells

Lookup NU author(s): Dr Zoltan Rakonczay, Dr Amy Fearn, Dr Michael Gray, Professor Barry Argent

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Abstract

Aim: To characterize H+ and HCO3- transporters in polarized CFPAC-1 human pancreatic duct cells, which were derived from a cystic fibrosis patient with the ΔF508 CFTR mutation. Methods: CFPAC-1 cells were seeded at high density onto permeable supports and grown to confluence. The cells were loaded with the pH-sensitive fluorescent dye BCECF, and mounted into a perfusion chamber, which allowed the simultaneous perfusion of the basolateral and apical membranes. Transmembrane base flux was calculated from the changes in intracellular pH and the buffering capacity of the cells. Results: Our results showed differential permeability to HCO3-/CO2 at the apical and basolateral membranes of CFPAC-1 cells. Na+/HCO3- co-transporters (NBCs) and Cl-/HCO3- exchangers (AEs) were present on the basolateral membrane, and Na+/H+ exchangers (NHEs) on both the apical and basolateral membranes of the cells. Basolateral HCO3- uptake was sensitive to variations of extracellular K+ concentration, the membrane permeable carbonic anhydrase (CA) inhibitors acetazolamide (100 μmol/L) and ethoxyzolamide (100 μmol/L), and was partially inhibited by H2-DIDS (600 μmol/L). The membrane-impermeable CA inhibitor 1-N-(4-sulfamoylphenylethyl)-2,4,6-trimethylpyridine perchlorate did not have any effect on HCO3- uptake. The basolateral AE had a much higher activity than that in the apical membrane, whereas there was no such difference with the NHE under resting conditions. Also, 10 μmol/L forskolin did not significantly influence Cl-/HCO3- exchange on the apical and basolateral membranes. The administration of 250 μmol/L H2-DIDS significantly inhibited the basolateral AE. Amiloride (300 μmol/L) completely inhibited NHEs on both membranes of the cells. RT PCR revealed the expression of pNBC1, AE2, and NHE1 mRNA. Conclusion: These data suggest that apart from the lack of CFTR and apical Cl-/HCO3- exchanger activity, CFPAC-1 cells express similar H+ and HCO3- transporters to those observed in native animal tissue. © 2006 The WJG Press. All rights reserved.


Publication metadata

Author(s): Rakonczay Jr Z, Fearn A, Hegyi P, Boros I, Gray MA, Argent BE

Publication type: Article

Publication status: Published

Journal: World Journal of Gastroenterology

Year: 2006

Volume: 12

Issue: 6

Pages: 885-895

ISSN (print): 1007-9327

ISSN (electronic): 1009-3079

Publisher: WJG Press

PubMed id: 16521216


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