Basic Research
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World J Gastroenterol. Feb 14, 2006; 12(6): 885-895
Published online Feb 14, 2006. doi: 10.3748/wjg.v12.i6.885
Characterization of H+ and HCO3- transporters in CFPAC-1 human pancreatic duct cells
Zoltán Rakonczay Jr, Amy Fearn, Péter Hegyi, Imre Boros, Michael A Gray, Barry E Argent
Zoltán Rakonczay Jr, Amy Fearn, Péter Hegyi, Michael A Gray, Barry E Argent, Institute for Cell and Molecular Biosciences, University of Newcastle upon Tyne, Medical School, Newcastle upon Tyne NE2 4HH, United Kingdom
Zoltán Rakonczay Jr, Péter Hegyi, First Department of Medicine, Faculty of Medicine, University of Szeged, H-6720 Szeged, Hungary
Imre Boros, Hungarian Academy of Sciences, Biological Research Center, Institute of Biochemistry, H-6726 Szeged, Hungary
Supported by a Wellcome Trust Travelling Fellowship to Z.R., No. 069470, and a Wellcome Trust IRDA Grant to P.H., No. 068096
Correspondence to: Professor Barry E Argent, Institute for Cell and Molecular Biosciences, University of Newcastle upon Tyne, Medical School, Framlington Place, Newcastle upon Tyne NE2 4HH, United Kingdom. b.e.argent@ncl.ac.uk
Telephone: +44-191-222-7009 Fax: +44-191-222-7424
Received: July 18, 2005
Revised: July 25, 2005
Accepted: August 3, 2005
Published online: February 14, 2006
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.

Keywords: CFPAC-1, Pancreatic duct cells, HCO3-, Intracellular pH