The expression of retinal cell markers in human retinal pigment epithelial cells and their augmentation by the synthetic retinoid fenretinide

PURPOSE In several species the retinal pigment epithelium (RPE) has the potential to transdifferentiate into retinal cells to regenerate functional retinal tissue after injury. However, this capacity for regeneration is lost in mammals. The synthetic retinoic acid derivative, fenretinide [N(4-hydroxyphenyl) retinamide], induces a neuronal-like phenotype in the human adult retinal pigment epithelial cell line (ARPE-19). These changes are characterized by the appearance of neural-like processes and the expression of neuronal markers not normally associated with RPE cells. Here we assess whether fenretinide can induce a neuroretinal cell phenotype in ARPE-19 cells, by examining retinal cell marker expression. METHODS ARPE-19 cells were treated daily with culture medium containing either 3 μM fenretinide or dimethyl sulfoxide as a control for 7 days. Cells were processed for immunocytochemistry, western blotting, and for analysis by PCR to examine the expression of a panel of RPE, neural, and retinal-associated cellular markers, including classical and non-canonical opsins. RESULTS Treatment with fenretinide for 7 days induced the formation of neuronal-like processes in ARPE-19 cells. Fenretinide induced the expression of the cone long wavelength sensitive opsin (OPN1lw) but not rhodopsin (RHO), while decreasing the expression of RPE cell markers. Many of the neuronal and retinal specific markers examined were expressed in both control and fenretinide treated cells, including those involved in photoreceptor cell development and the multipotency of neural retinal progenitor cells. Interestingly, ARPE-19 cells also expressed both photoreceptor specific and non-specific canonical opsins. CONCLUSIONS The expression of retinal-associated markers and loss of RPE cell markers in control ARPE-19 cells suggests that these cells might have dedifferentiated from an RPE cell phenotype under standard culture conditions. The expression of molecules, such as the transcription factors paired box 6 gene (PAX6), sex determining region Y-box 2 (SOX2), cone-rod homeobox (CRX), and neural retina leucine zipper (NRL), further implies that in culture these cells are predisposed toward a retinal progenitor-like state. The fenretinide-induced increase in photoreceptor cell markers, accompanied by a decrease in RPE cell markers, suggests that retinoids may play a role in the transdifferentiation of RPE cells. Importantly, our data show for the first time the expression of a vertebrate ciliary opsin (OPN1lw) and rhabdomeric-like opsin, opsin 4 (OPN4 also known as melanopsin) in a clonal cell line. Together these data suggest that ARPE-19 cells are primed for and possess the capacity to differentiate toward a retinal cell-like lineage.