The Core uses transepithelial electrophysiological measurements of confluent cultured monolayers, as well as native tissue, mounted in Ussing chambers to determine the magnitude of both Cl- (CFTR) and Na+ (ENaC) currents. Fluid transport rates are measured in vitro by both the blue dextran technique and by confocal microscopy of fluorescently labeled ASL.
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Analysis of efficacy of CFTR gene transfer to CF human and murine airways
In Vitro
1) Measure ion transport properties of “corrected” vs. control CF human and murine airway cultures in Ussing chambers.
2) Measure fluid transport properties of “corrected” vs. control CF human and murine airway cultures by confocal microscopy.Measure fluid transport properties of “corrected” vs. control CF human and murine airway cultures by blue dextran optical absorption.
3) Measure pH from the lumenal surface of well-differentiated “corrected” vs. control CF human and murine airway cultures.
4) Measure mucus biophysical properties and mucus transport in corrected vs control CF airway cultures.
5) Measure ion transport properties of freshly excised murine neonatal airways of mice corrected inin vivo vs control mice.
In Vivo
1) Measure electrical potential difference (PD) in murine nasal and nasopharyngeal epithelia in vivo.
2) Measure nasal PD in humans.
Analysis of efficacy of CFTR gene transfer to gastrointestinal epithelium of both mouse (neonatal and adult) and human
1) Measure ion transport properties of freshly-excised murine neonatal and adult gut epithelia as well as transit times in CF corrected vs non-corrected gut epithelia.
2) Measure ion transport properties of rectal biopsies from human CF patients.
Analysis of efficacy of functional and biochemical correction of mutant CFTR by small-molecule compounds.
1) Measure ion transport properties of "corrected" vs. control CF human airways cultures in Ussing chambers.
2) Quantitate CFTR maturation by Western blotting and analyze CFTR cell surface localization by On-Cell Westerns.