CFTR upregulates the expression of the basolateral Na+-K+-2Cl- cotransporter in cultured pancreatic duct cells

The purpose of the current experiments was 1) to assess basolateral Na+-K+-2Cl(-) cotransporter (NKCC1) expression and 2) to ascertain the role of cystic fibrosis transmembrane conductance regulator (CFTR) in the regulation of this transporter in a prototypical pancreatic duct epithelial cell Line. Previously validated human pancreatic duct cell Lines (CFPAC-1), which exhibit physiological features prototypical of cystic fibrosis, and normal pancreatic duct epithelia (stable recombinant CFTR-bearing CFPAC-1 cells, termed CFPAC-WT) were grown to confluence before molecular and functional studies. High-stringency Not-them blot hybridization, utilizing specific cDNA probes, confirmed that NKCC1 was expressed in both cell lines and its mRNA levels were twofold higher in CFPAC-WT cells than in CFPAC-1 cells (P < 0.01, n = 3). Na+-K+-2Cl(-) cotransporter activity, assayed as the bumetanide-sensitive, Na+- and Cl--dependent NH4+ entry into the cell (with NH4+ acting as a substitute for K+), increased by similar to 115% in CFPAC-WT cells compared with CFPAC-1 cells (P < 0.01, n = 6). Reducing the intracellular Cl- by incubating the cells in a Cl--free medium increased Na+-K+-2Cl(-) cotransporter activity by twofold (P < 0.01, n = 4) only in CFPAC-WT cells. We concluded that NKCC1 is expressed in pancreatic duct cells and mediates the entry of Cl-. NKCC1 activity is enhanced in the presence of an inward Cl- gradient. The results further indicate that the presence of functional CFTR enhances the expression of NKCC1. We speculate that CFTR regulates this process in a Cl--dependent manner.