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


METHODS
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.


RESULTS
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.


CONCLUSIONS
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.
Krugluger
Walter
W
Department of Clinical Chemistry, The Ludwig Boltzmann Institut of Retinology and Biomicroscopic Lasersurgery, Rudolfstiftung Hospital, Vienna, Austria.
Seidel
Stefan
S
Steindl
Kerstin
K
Binder
Susanne
S
D002352:Carrier Proteins / Q000235:genetics; D002460:Cell Line; D049109:Cell Proliferation / Q000187:drug effects; D004815:Epidermal Growth Factor / Q000494:pharmacology*; D005136:Eye Proteins / Q000235:genetics; D038362:Glycogen Synthase Kinase 3 / Q000235:genetics*; D006801:Humans; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D053538:Keratin-18 / Q000235:genetics; D010857:Pigment Epithelium of Eye / Q000187:drug effects* / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D011994:Recombinant Proteins / Q000494:pharmacology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014158:Transcription, Genetic / Q000187:drug effects*; D051176:beta Catenin / Q000235:genetics*; D019745:cis-trans-Isomerases; D000099178:Retinoid Isomerohydrolase
D002352:Carrier Proteins; D004815:Epidermal Growth Factor; D005136:Eye Proteins; D038362:Glycogen Synthase Kinase 3; D007334:Insulin-Like Growth Factor I; D053538:Keratin-18; D012333:RNA, Messenger; D011994:Recombinant Proteins; D051176:beta Catenin; D019745:cis-trans-Isomerases; D000099178:Retinoid Isomerohydrolase
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