CFTR channels in immortalized human airway cells

Abstract

The cystic fibrosis (CF) gene codes for CF transmembrane regulator (CFTR), a small-conductance linear Cl- channel, but numerous studies have identified a larger conductance, rectifying Cl- channel as the adenosine 3',5'-cyclic monophosphate (cAMP)-regulated channel that is defective in airway cells. We examined Cl- conductance in a bronchial epithelial cell line that expresses CFTR, 16HBE14o-, (CFTR+) and in an airway cell line that does not, 9HTEo-/S, (CFTR-). Ionomycin or hypotonic Ringer increased iodide efflux from both cell lines; however, forskolin increased iodide efflux or whole cell Cl- currents only in CFTR+ cells. Forskolin-stimulated whole cell currents were linear, voltage independent, and blocked by iodide. Cell-attached and outside-out patches from confluent CFTR+ but not CFTR- cells revealed 6-pS channels having linear current-voltage relations, permselectivity Cl > I (partial block by external iodide), and little or no inhibition by 5-nitro-2-(3-phenylpropylamino)-benzoate. The number of active channels per patch increased from 0.6 to 3.0 after forskolin. Channels closed after excision with tau = 4 s, but activity could be prolonged with ATP or protein kinase A plus ATP. Channels were modeled with one open and four closed states and show apparent cooperativity in gating. Rectifying Cl- channels previously implicated in CF were not seen in cell-attached recordings from either cell line but were abundant in excised patches from both cell lines. Thus CFTR channels are the pathway for cAMP-mediated Cl- conductance in these human airway cells, Ca2+ and swelling-induced channels do not require CFTR, and CFTR-cells display a CF phenotype.

Characterization of cAMP dependent CFTR‐chloride channels in human trache gland cells

“…Recently, using immortalized human airway cell lines, Haws et al [18] identified a linear 6 pS Cl-channel activated by forskolin in cell-attached patches. This channel has properties closely related to those of the channel described in this paper.…”

“…CF causes a decrease in CAMP-dependent [ 14,151 but not calciumdependent regulation of chloride transport in the airway [14]. Various Cl-channels have been identified in tracheal surface epithelial cells [ 16,171, including CFTRCl-channels in immortalized human airway cell lines [18] but there have been no single channel studies reported for submucosal gland cells, although these cells are believed to be the major site of mucus secretion in the airway epithelium in situ hybridization and immunocytochemistry indicate that CFTR expression is far higher in submucosal glands than in other airway cells [21]. Until now, CFTR chloride channels have not been described in human pulmonary cells from submucosal gland cells.…”

Anion efflux from cytotrophoblast cells derived from normal term human placenta is stimulated by hyposmotic challenge and extracellular A23187 but not by membrane‐soluble cAMP

“…The unidirectional non-equilibrium efflux of radioactive ions from cells has been used as a marker of ion transport and secretion in several contexts (Venglarik et al 1990), for example (Clancy et al 1990;Haws et al 1992).…”

Cystic Fibrosis: Does CFTR Malfunction Alter pH Regulation?

“…In 1991 the CFTR channel was confirmed as a small linear chloride channel by Bear et al when they expressed recombinant CFTR in cells [9]. Once the small CFTR channels identity was confirmed Wine and Krouse teamed with Christine Haws to be the first to describe a kinetic model for the gating of CFTR [88]. This model has since been modified and expanded, by researchers such as David Sheppard in the UK and TC Hwang in Missouri, but it was the first.…”