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S. Tamiya, L. Liu, H. J. Kaplan, D. C. Dean; Epithelial-to-Mesenchymal Transition in RPE Cells - A Switch in Cadherin Expression From P-Cadherin to N-Cadherin. Invest. Ophthalmol. Vis. Sci. 2009;50(13):2702.
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Retinal pigment epithelial (RPE) cell proliferation and epithelial-to-mesenchymal transition (EMT) have been implicated to play a role in the development of proliferative vitreoretinopathy (PVR). Our in vitro model of PVR using sheets of RPE in primary culture shows that proliferation and EMT are linked, and that these processes are restricted to cells at the edge of the RPE sheets where cell-cell contacts are lost. The purpose of this study was to examine the change in expression of cell adhesion molecules during RPE cell proliferation and EMT.
RPE cell sheets, isolated from porcine eyes using Dispase, were cultured on porcine lens posterior capsule for up to 8 days in DMEM supplemented with fetal bovine serum. RPE cells with their underlying lens capsule were micro-dissected, lysed and used for western blot analyses. In addition, RPE sheets were fixed at various time points in culture for immunohistochemical staining of cell-cell adhesion molecules (E-, N-, and P-cadherin as well as ZO-1) or markers of proliferation or EMT (BrdU and vimentin, respectively).
RPE cells at the edge of the sheets initiated proliferation, whereas cells retaining cell-cell contact in the interior of the sheets remained non-proliferative. Concomitant with proliferation, cells at the edge of the sheets underwent EMT, losing their cuboidal epithelial morphology and pigmentation. Western blot analyses revealed a significant switch in cadherin expression from P-cadherin to N-cadherin in the cells undergoing EMT. Immunostaining of the cultured cells confirmed the findings from western blots and demonstrated that N-cadherin replaces P-cadherin at cell-cell borders as RPE cells acquire fibroblastic morphology. In contrast to the switch in cadherin expression, ZO-1 was maintained at cell-cell borders during EMT.
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