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D. Yu, W. Thelin, T. Rogers, S. Randell, B. Grubb, R. Boucher; Conjunctival Ion Transport in Cystic Fibrosis and Control Mice. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1909.
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Adequate electrolyte and concomitant liquid transport is essential for conjunctival epithelial health. Trans-cellular ion transport drives osmotic water flux that either decreases the aqueous tear film (sodium absorption) or increases ocular hydration (chloride secretion). The objective of the current study was to investigate ion transport across cystic fibrosis and normal mouse conjunctiva.
Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR) was performed to characterize epithelial sodium channel (ENaC), cystic fibrosis transmembrane conductance regulator (Cftr), and calcium activated chloride channel (Tmem16a) mRNA expression in mouse conjunctiva. Conjunctival tissues of Cftr knockout (Cftr -/-) mice and heterozygous littermate control (Cftr +/-) mice were mounted on Ussing chambers for measurement of electrophysiological properties. Conjunctival histology of both mouse groups was evaluated.
RT-PCR demonstrated Cftr, ENaC (α, β, and γ subunits), and Tmem16a mRNA expression in mouse conjunctiva. In Ussing chambers, the conjunctival basal short circuit current (Isc) of Cftr +/- mice (38.38 ± 5.26 µA, n=11) was significantly higher than the Isc of Cftr -/- mice (15.23 ± 4.18 µA, n=11) (P<0.01). Addition of the CFTR inhibitor-172 produced a decrease in Isc of 19.07 ± 3.27 µA (n=11) in Cftr +/- littermate control mice and a decrease in Isc of 1.89 ± 0.58 µA (n=11) in Cftr -/- mice (P<0.01). In additional preparations, forskolin (CFTR activator) stimulated a ΔIsc of 59.21 ± 7.40 µA (n=10) in control mice vs. a ΔIsc of 6.34 ± 1.37 µA (n=12) in knockout mice (P<0.05), respectively. Inhibition of sodium transport with amiloride (ENaC inhibitor) or phloridzin (Sodium/glucose cotransporter inhibitor) produced similar but small inhibitory effects on Isc in both mouse groups. There were no gross phenotypic changes in Cftr knockout mouse conjunctiva and histology did not reveal changes in fine structure including epithelial goblet cell density or inflammation.
These studies indicate that Cftr dominates the ion transport phenotype of the normal mouse conjunctiva. In Cftr knockout mice, Tmem16a may compensate for Cftr to mediate sufficient chloride secretion across the conjunctiva to prevent the development of overt pathology.
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