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J. Biol. Chem., Vol. 265, Issue 22, 12796-12800, Aug, 1990

A cystic fibrosis phenotype in cells cultured from sweat gland secretory coil. Altered kinetics of 36Cl efflux

LC Wood and EF Neufeld
Department of Biological Chemistry, School of Medicine, University of California, Los Angeles 90024.

As a step toward understanding the metabolic consequences of the cystic fibrosis (CF) mutation, we have examined the kinetics of 36Cl efflux in cells cultured from sweat glands, a tissue that is affected in the disease. Epithelial cells, derived from the secretory coil of sweat glands of CF and control individuals, were cultured in serum-free medium, and primary cultures used for efflux experiments. Cell layers were equilibrated with Na36Cl in 4-(2-hydroxyethyl)-1- piperazineethanesulfonic acid-buffered balanced salt solution for 45 min at 37 degrees C, washed in 0.25 M sucrose, and incubated in nonradioactive buffer for measurement of 36Cl efflux. Efflux from CF and control cells followed biphasic kinetics and was described by the equation Y = Ae-kat + Be-kbt. All efflux was inhibited at 6 degrees C. The fast component of efflux, Ae-kat, of both control and CF cells was inhibited by the anion channel blockers 4,4'-diisothiocyanato-2,2'- stilbene disulfonic acid, 9-anthracene carboxylate, and diphenylamine 2- carboxylate, implicating release through chloride channels. At 23 degrees C, the kinetics of 36Cl efflux from CF and control cells were indistinguishable, but efflux from control cells could be accelerated by cAMP analogs and isoproterenol. At 37 degrees C, 36Cl efflux was more rapid from control cells than from CF cells, but could not be stimulated further by beta-adrenergic agents. In both cases, the increased rate of efflux was due to a severalfold increase in the A parameter of the fast component. These differential responses constitute a "CF phenotype" of secretory sweat gland cells in culture that may be useful for further investigation of the metabolic defect in cystic fibrosis.
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