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Lens epithelium and fiber Na,K-ATPases: distribution and localization by immunocytochemistry.
Invest Ophthalmol Vis Sci. 1999 Sep; 40(10):2291-8.IO

Abstract

PURPOSE

To use immunofluorescence and immunogold techniques to identify the catalytic subunits of the Na,K-ATPases of the lens and to determine their location in the cells of the epithelium and cortex of bovine and human lenses.

METHODS

Frozen sections of capsulated and decapsulated bovine and human lenses were prepared, blocked, and treated with affinity-purified polyclonal rabbit antibodies to the Na,K-ATPase catalytic subunit isoforms with subsequent treatment with fluorescein isothiocyanate-labeled goat anti-rabbit IgG and visualization of the fluorescence by light microscopy. An immunogold-labeled goat anti-rabbit IgG was used to detect, by electron microscopy, the binding of the same affinity-purified polyclonal antibodies to thin sections of decapsulated lenses that had been fixed and embedded in Lowicryl K4M. The results were confirmed by staining of western blot analysis of sodium dodecyl sulfate-polyacrylamide gel separations of enriched membrane preparations from bovine and human lenses.

RESULTS

The three common catalytic subunits of the Na,K-ATPases are present in the plasma membranes of lens epithelium, lens fibers, or both. The data indicate a polarized distribution of the alpha1 and alpha3 catalytic subunit isoforms in central epithelium. In the cortical fibers, the alpha2 isoform is present around the interdigitations. The alpha3 isoform is found in the interdigitation-free regions of human cortical fibers.

CONCLUSIONS

This unique distribution of Na,K-ATPases precludes the popular pump-leak model for lens monovalent cation homeostasis. The functional significance of the distribution of Na,K-ATPases in the lens epithelium and superficial fibers is currently under investigation.

Authors+Show Affiliations

University of North Texas Health Science Center, Department of Anatomy and Cell Biology, Fort Worth 76107, USA.No affiliation info available

Pub Type(s)

Journal Article
Research Support, U.S. Gov't, P.H.S.

Language

eng

PubMed ID

10476795

Citation

Garner, M H., and Y Kong. "Lens Epithelium and Fiber Na,K-ATPases: Distribution and Localization By Immunocytochemistry." Investigative Ophthalmology & Visual Science, vol. 40, no. 10, 1999, pp. 2291-8.
Garner MH, Kong Y. Lens epithelium and fiber Na,K-ATPases: distribution and localization by immunocytochemistry. Invest Ophthalmol Vis Sci. 1999;40(10):2291-8.
Garner, M. H., & Kong, Y. (1999). Lens epithelium and fiber Na,K-ATPases: distribution and localization by immunocytochemistry. Investigative Ophthalmology & Visual Science, 40(10), 2291-8.
Garner MH, Kong Y. Lens Epithelium and Fiber Na,K-ATPases: Distribution and Localization By Immunocytochemistry. Invest Ophthalmol Vis Sci. 1999;40(10):2291-8. PubMed PMID: 10476795.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Lens epithelium and fiber Na,K-ATPases: distribution and localization by immunocytochemistry. AU - Garner,M H, AU - Kong,Y, PY - 1999/9/7/pubmed PY - 1999/9/7/medline PY - 1999/9/7/entrez SP - 2291 EP - 8 JF - Investigative ophthalmology & visual science JO - Invest Ophthalmol Vis Sci VL - 40 IS - 10 N2 - PURPOSE: To use immunofluorescence and immunogold techniques to identify the catalytic subunits of the Na,K-ATPases of the lens and to determine their location in the cells of the epithelium and cortex of bovine and human lenses. METHODS: Frozen sections of capsulated and decapsulated bovine and human lenses were prepared, blocked, and treated with affinity-purified polyclonal rabbit antibodies to the Na,K-ATPase catalytic subunit isoforms with subsequent treatment with fluorescein isothiocyanate-labeled goat anti-rabbit IgG and visualization of the fluorescence by light microscopy. An immunogold-labeled goat anti-rabbit IgG was used to detect, by electron microscopy, the binding of the same affinity-purified polyclonal antibodies to thin sections of decapsulated lenses that had been fixed and embedded in Lowicryl K4M. The results were confirmed by staining of western blot analysis of sodium dodecyl sulfate-polyacrylamide gel separations of enriched membrane preparations from bovine and human lenses. RESULTS: The three common catalytic subunits of the Na,K-ATPases are present in the plasma membranes of lens epithelium, lens fibers, or both. The data indicate a polarized distribution of the alpha1 and alpha3 catalytic subunit isoforms in central epithelium. In the cortical fibers, the alpha2 isoform is present around the interdigitations. The alpha3 isoform is found in the interdigitation-free regions of human cortical fibers. CONCLUSIONS: This unique distribution of Na,K-ATPases precludes the popular pump-leak model for lens monovalent cation homeostasis. The functional significance of the distribution of Na,K-ATPases in the lens epithelium and superficial fibers is currently under investigation. SN - 0146-0404 UR - https://www.unboundmedicine.com/medline/citation/10476795/Lens_epithelium_and_fiber_NaK_ATPases:_distribution_and_localization_by_immunocytochemistry_ L2 - https://iovs.arvojournals.org/article.aspx?volume=40&issue=10&page=2291 DB - PRIME DP - Unbound Medicine ER -