Considerable progress has been made in CF research since the demonstration of low Cl- permeability in epithelia of target organs. Recent studies by others on cultured tracheal epithelial cells indicate that the defect in Cl- transport is not in the Cl- channels, but a malfunction in regulation of the Cl- pathway. Advances at the molecular level have been no less exciting. Linkage studies have localized the CF gene to chromosome 7q22-31 and a candidate for the cystic fibrosis locus has now been isolated. Identification of the CF gene and its product is only a beginning. Definition of the metabolic defect require an understanding of the abnormal gene product and its physiological expression. The goal of the research in the proposed project is to establish immortal cell lines from the human eccrine sweat gland for use as a model system for studying the basic physiological and biochemical defects in CF. The human eccrine sweat gland is an appropriate model for CF research, since sweat glands from CF subjects: i) demonstrate defects in secretory and reabsorptive functions ii) appear to express differences in the phosphorylation of a specific polypeptide iii) are not burdened by other pathological manifestations; and iv) are relatively accessible. Establishing cultured cell lines from the sweat gland will increase the quantity and longevity of the tissue and provide the opportunity to study and compare the two regions of the sweat gland (the secretory coil and reabsorptive duct) independently and in a simplified model. We propose to characterize the cells in culture to determine the extent to which they resemble the progenitor tissue, and more importantly to demonstrate the persistence of the phenotypic expression of the CF genotype in the cultured cells. Specifically, we will perform electrophysiological studies of Cl- conductance its regulation by various agonists as well as analyses of phosphorylation profiles of polypeptides. Expression of the characteristic CF phenotype in culture will allow us to investigate the underlying metabolic defect. Eventually we hope to be able to alter its expression toward a therapeutic control of CF pathology.
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