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S. Tamiya, Q. Z. Ruley, H. J. Kaplan, D. C. Dean; Lithium Chloride Prevents Ectopic Proliferation and Dedifferentiation of RPE Cells. Invest. Ophthalmol. Vis. Sci. 2008;49(13):5557.
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Various studies have shown that ectopic proliferation and dedifferentiation of retinal pigment epithelial (RPE) cells to a mesenchymal phenotype leads to formation of scar-like tissue following retinal detachment which is the major cause of retinal reattachment surgery failure in proliferative vitreoretinopathy (PVR). In this study, an in vitro culture model was utilized to examine the early changes occurring to RPE cells and to potentially modify these changes.
Following removal of the sclera, porcine eyes were digested with dispase and RPE cells were isolated as cell sheets. Harvested RPE sheets were cultured on porcine lens posterior capsule in DMEM supplemented with fetal bovine serum. In some cultures, 20mM LiCl, a GSK-3β inhibitor which has been shown to modify cell growth in other cell types, or NaCl (used as a control) were added to the culture media after the initial cell attachment. BrdU uptake was utilized to examine cell proliferation. RPE cell dedifferentiation was examined by immunohistochemical staining of vimentin, a mesenchymal marker that is classically induced upon epithelial-mesenchymal transition.
RPE sheets were able to attach to the posterior capsule within 6 hours and migration of the cells at the edge of the sheet were observed within 30 hours. BrdU staining revealed proliferation of RPE cells, mainly at the edge of the sheet, as early as 2 days. These migrating and proliferating cells became larger in size, less pigmented and stained positive for vimentin. In contrast, confluent cells at the center of sheet remained heavily pigmented and retained their epithelial morphology with little proliferation or vimentin expression. Addition of LiCl to the culture media significantly inhibited RPE cell migration and proliferation. In the presence of LiCl, the majority of the cells remained heavily pigmented and the number of BrdU positive cells was significantly less compared to the NaCl control. Furthermore, the number of vimentin-positive cells was significantly reduced in the presence of LiCl.
RPE cells in our in vitro culture model displayed the two major changes, ectopic cell proliferation and dedifferentiation, which have been shown to play a major role in PVR development. Both changes were limited to the edge of the cell sheet implying that loss of cell-cell contacts, as cells migrate away from the other cells in the RPE sheet, triggers the ectopic proliferation and dedifferentiation. LiCl, a widely used GSK-3β inhibitor, significantly inhibited these changes suggesting that it could potentially be used for the treatment of PVR.
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