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M. Srinivas, Y. Gao, L. Li, C. Sellitto, T. W. White; Changes in Lens Epithelial Cell Proliferation Are Correlated With Changes in Gap Junctional Coupling. Invest. Ophthalmol. Vis. Sci. 2007;48(13):3634.
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© ARVO (1962-2015); The Authors (2016-present)
Gap junctions between epithelial cells are essential for normal lens growth. In mice, knockout of Cx50 or targeted replacement of Cx50 with Cx46 (knockin) causes smaller lenses due to decreased epithelial cell proliferation. However, it remains unclear whether Cx50 functionally contributes to lens epithelial coupling during maximal mitosis on postnatal days 2 and 3 (P2-P3). To determine which connexins functionally contribute to epithelial cell coupling and proliferation, we examined junctional coupling from epithelial cells of wild-type and knockin mice.
Epithelial cells were isolated from wild-type or knockin mice at different developmental ages. Junctional currents were measured by dual whole cell voltage clamp. Cell proliferation was assayed by BrdU incorporation. Connexins were immunolocalized using specific antibodies.
Junctional currents between lens epithelial cells were strongly sensitive to quinine, a drug that blocks Cx50 gap junctions, but not Cx43 or Cx46. Single channel currents had a unitary conductance of 210 pS, typical of Cx50. Immunocytochemical staining showed Cx43 and Cx50 were abundantly expressed in wild-type cells and Cx46 replaced Cx50 in knockin cells. A correlation between functional activity of Cx50 and maximal mitosis was also found. In epithelial cells from P3 mice, there was a high incidence of clusters of proliferating epithelial cells in wild-type mice and >60% of total coupling was provided by Cx50. In adult cells, mitosis was greatly reduced and Cx50 contributed ~30% of the total coupling. Cx46 knockin mice which displayed fewer clusters of proliferating cells on P3, and coupling between cells was similar in magnitude to wild-type, but had pharmacological and biophysical characteristics of Cx46.
We have shown that Cx50 is abundantly expressed and functionally active in lens epithelial cells at the time of maximal proliferation and that Cx46 could not substitute for Cx50. These results suggest that the developmentally regulated proliferation of epithelial cells requires the unique functional activity of Cx50.
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