Epidermal growth factor inhibits glycogen synthase kinase-3 (GSK-3) and β-catenin transcription in cultured ARPE-19 cells

Abstract

Culture of retinal pigment epithelium (RPE) cells might be a future option in the therapy of various degenerative retinal diseases. However, the molecular changes which occur during in vitro expansion of RPE cells during culture are not fully elucidated. The aim of this study was to evaluate molecular changes in the RPE cell line ARPE-19 after stimulation with different growth factors. Cultured ARPE-19 cells were stimulated for 72 hours with rh-EGF, rh-IGF-1, rh-VEGF or rh-bFGF, and transcriptional changes of the differentiation markers cytokeratin 18 and RPE65 and of the key molecules of the wnt pathway, beta-catenin, and glycogen synthase kinase-3 (GSK-3) were evaluated by real time RT-PCR. We found a significant decrease of cytokeratin 18 and RPE65 transcription after stimulation with rh-EGF (0.47 +/- 0.42 and 0.32 +/- 0.57-fold, respectively; p < 0.05). A significant reduction of beta-catenin and GSK-3 mRNA was found in ARPE-19 cells stimulated with rh-IGF-1 (0.61 +/- 0.25 and 0.52 +/- 0.02-fold, respectively) or rh-EGF (0.55 +/- 0.19 and 0.76 +/- 0.26-fold, respectively). No changes of beta-catenin mRNA were observed after stimulation with rh-VEGF or bFGF. Our data suggest an inhibition of the beta-catenin-pathway in ARPE-19 cells by IGF-1 and EGF, suggesting that ARPE-19 cell proliferation is, at least in part, driven by the beta-catenin pathway. Furthermore, induction of proliferation by EGF results in a loss of differentiation markers in these cells. Maintaining the RPE phenotype is still one of the main problems for RPE- transplantation.