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An immortal cell line to study the role of endogenous CFTR in electrolyte absorption
- Source :
- In vitro cellulardevelopmental biology. Animal. 31(1)
- Publication Year :
- 1995
-
Abstract
- The intact human reabsorptive sweat duct (RD) has been a reliable model for investigations of the functional role of “endogenous” CFTR (cystic fibrosis transmembrane conductance regulator) in normal and abnormal electrolyte absorptive function. But to overcome the limitations imposed by the use of fresh, intact tissue, we transformed cultured RD cells using the chimeric virus Ad5/SV40 1613 ori-. The resultant cell line, RD2(NL), has remained differentiated forming a polarized epithelium that expressed two fundamental components of absorption, a cAMP activated Cl− conductance (Gcl) and an amiloride-sensitive Na+ conductance (GNa). In the unstimulated state, there was a low level of transport activity; however, addition of forskolin (10−5 M) significantly increased the Cl− diffusion potential (Vt) generated by a luminally directed Cl− gradient from − 15.3 ± 0.7 mV to −23.9 ± 1.1 mV,n=39; and decreased the transepithelial resistance (Rt) from 814.8 ± 56.3 Ω.cm2 to 750.5 ± 47.5 Ω.cm2,n=39, (n=number of cultures). cAMP activation, anion selectivity (Cl−>I−>gluconate), and a dependence upon metabolic energy (metabolic poisoning inhibited GCl), all indicate that the GCl expressed in RD2(NL) is in fact CFTR-GCl. The presence of an apical amiloride-sensitive GNa was shown by the amiloride (10−5 M) inhibition of GNa as indicated by a reduction of Vt and equivalent short circuit current by 78.0 ± 3.1% and 77.9 ± 2.6%, respectively, and an increase in Rt by 7.2 ± 0.8%,n=36. In conclusion, the RD2(NL) cell line presents the first model system in which CFTR-GCl is expressed in a purely absorptive tissue. It provides an opportunity to study the properties and role of CFTR in the context of absorptive function in immortalized epithelial cells.
- Subjects :
- Adult
Anions
medicine.medical_specialty
Cystic Fibrosis Transmembrane Conductance Regulator
Endogeny
Context (language use)
Eccrine Glands
Epithelium
Absorption
Amiloride
chemistry.chemical_compound
Chlorides
Internal medicine
medicine
Cyclic AMP
Humans
Cell Line, Transformed
Forskolin
biology
Colforsin
Sodium
Electric Conductivity
Membrane Proteins
Epithelial Cells
Cell Biology
General Medicine
Molecular biology
Cystic fibrosis transmembrane conductance regulator
Endocrinology
medicine.anatomical_structure
chemistry
Cell culture
biology.protein
Female
Immortalised cell line
Developmental Biology
medicine.drug
Subjects
Details
- ISSN :
- 10712690
- Volume :
- 31
- Issue :
- 1
- Database :
- OpenAIRE
- Journal :
- In vitro cellulardevelopmental biology. Animal
- Accession number :
- edsair.doi.dedup.....0dfffa65e580eb67ce3942ba00c21ae2