Vitreous-induced Transformation of Human RPE Cells Involves Changes in the Expression of Specific Transcription Factors

Abstract

Abstract: : Purpose: When human RPE cells come in contact with vitreous, they undergo an epithelial-mesenchymal transformation. This is accompanied by alterations in the expression of a number of genes, several of which contain promotor binding sites for the transcription factors Egr-1 and AP-1. Investigations were carried out to determine whether RPE cell transformation is accompanied by changes in expression and/or location of these transcription factors. Methods: Human RPE cells were grown in control medium or medium supplemented with 25% human vitreous. Changes in gene expression were monitored over a 48-hour period using gene arrays and were confirmed by real-time PCR. Changes in protein expression were measured by western blotting. The location of proteins was determined by confocal microscopy. Results: Vitreous-induced transformation is accompanied by an increase in the transcription of a number of mRNAs including those encoding prostaglandin E2 synthase, COX-2, heme oxygenase-1, metalloproteinase-14 and VEGF. Also up-regulated is c-Fos. The latter forms a complex with c-Jun that associates with the AP-1 binding site. In addition, Egr-1, which is found in the cytoplasm of untransformed RPE cells, migrates to the nucleus on vitreous-induced transformation. Conclusions: Transformation of human RPE cells is accompanied by the changes in a variety of proteins including growth factors, ECM receptors and metalloproteinases. The transcription of some of these proteins may be under the control of Egr-1 and c-Fos, two transcription factors whose behavior is also altered during transformation.

Keywords: retinal pigment epithelium • vitreous • transcription factors