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Dongfang Yu, William Thelin, Troy Rogers, Scott Randell, Richard Boucher; Ion Transport Characterization of the Rat Lacrimal Functional Unit. Invest. Ophthalmol. Vis. Sci. 2011;52(14):3707.
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Active ion transport and coupled osmotic water flow is essential for the maintenance of ocular surface health. The present study was designed to investigate the role of specific sodium and chloride channels in the ion transport activities of selected ocular epithelia, including epithelia from conjunctiva, cornea, lacrimal gland, and Meibomian gland.
In rat ocular tissues, mRNA expression of the epithelial sodium channel (ENaC), sodium glucose co-transporter (Slc5a1), cystic fibrosis transmembrane conductance regulator (Cftr), calcium activated chloride channel (Tmem16a), and aquaporin (Aqp) 3 and 5 was determined by Reverse Transcriptase-Polymerase Chain Reaction (RT-PCR). Protein expression and localization of β-ENaC were evaluated by western blot and immunohistochemistry in ocular tissues. Freshly isolated conjunctiva tissues were mounted in Ussing chambers to study bioelectric properties. Primary cultures derived from palpebral conjunctiva and Meibomian gland tissues were also subjected to electrophysiologic measurements in Ussing chambers.
ENaC (α, β, and γ subunits), Slc5a1, Cftr, Aqp3, and Aqp5 mRNA were expressed in rat conjunctiva, cornea, lacrimal gland, and Meibomian gland tissues. The expression levels of β and γ ENaC were significantly higher in Meibomian gland tissues relative to other tissues. Tmem16a mRNA was expressed in all tissues except cornea, with the lacrimal gland having the highest level. Aqp5 was most abundantly present in cornea among all tissues. Western blots detected a 95kD band of β-ENaC most abundantly in the Meibomian gland. Immunofluorescence also localized the distribution of β-ENaC to the cytosol of Meibomian gland acinar cells. Electrophysiologic studies of freshly isolated conjunctival tissues identified functional ENaC, Slc5a1, Cftr, and Tmem16a by the application of activators or inhibitors of these channels. Fornical conjunctiva tissue exhibited the most active ion transport under basal conditions among the three regions of conjunctiva. Ussing chamber studies of cell culture derived from palpebral conjunctiva and Meibomian gland revealed contributions of ENaC, Slc5a1, and Tmem16a to basal ion transport.
This study demonstrated the involvement of multiple ion and water channels in conjunctiva, cornea, lacrimal gland, and Meibomian gland tissues in the rat. Investigation of the roles of these channels on different ocular epithelia may lead to development of new therapeutic agents for ocular surface disorders.
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