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R.J. Stump, F.J. Lovicu, J.W. McAvoy; Lithium Chloride Inhibits Proliferation, Migration and TGF–beta–Induced Epithelial Mesenchymal Transition (EMT) in Lens Epithelial Explants . Invest. Ophthalmol. Vis. Sci. 2004;45(13):1717.
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© ARVO (1962-2015); The Authors (2016-present)
Purpose: Research in our laboratory indicates that the Wnt growth factor family has a role in maintaining the differentiation of lens epithelial cells. Lithium chloride (LiCl) has been shown to promote Wnt signaling through the ß–catenin pathway. This study aimed to assess the effects of LiCl on lens epithelial cells in explant culture. Methods: Lens epithelial explants were prepared from postnatal day 21 rats. To test the effects of LiCl on cell proliferation and cell migration, LiCl was added to explant cultures over a range of concentrations. For controls, similarly treated explants were set up in NaCl at the same final concentrations. For proliferation studies, explants were incubated with BrdU for 6 hours prior to harvesting. Cells that had incorporated BrdU were identified using peroxidase immunohistochemistry. For migration studies, cellular distribution in explants was monitored daily. To test the effects of LiCl on TGFß–induced EMT, explants were cultured with TGFß and various combinations of LiCl. Results: In the presence of 15–30 mM LiCl, BrdU–incorporation was significantly inhibited in explants compared with controls, indicating cell–cycle arrest. In addition, the presence of LiCl inhibited cell spreading/migration. The cells in LiCl–treated explants strongly expressed molecules associated with their tight junctions, including ZO–1 and ß–catenin. Immunofluorescent localisation of these molecules around the cell margins defined the hexagonal shape of the cells and their cobblestone–like packing arrangement. In contrast, in the controls, the cells rapidly spread/migrated to cover bare patches of lens capsule. In this case, ZO–1 and ß–catenin were not localised to the tight junctions and the cells were squamous, with many lamellipodia and filipodia. In explants treated with TGFß, the cells became spindle–shaped, taking on a fibro–/myofibroblastic morphology, as well as expressing alpha–smooth muscle actin; all features of TGFß–induced EMT. In the presence of LiCl, this TGFß–induced EMT was blocked, with lens epithelial cells retaining their cuboidal morphology and cobblestone–like packing arrangement. In addition, none of these cells expressed alpha–smooth muscle actin. Conclusions: LiCl is a very effective blocker of endogenous cell proliferation and cell spreading/migration in lens epithelial explants. LiCl also inhibits TGFß–induced EMT and maintains the lens epithelial phenotype in explant cultures.
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