Characterization of a Spontaneously Transformed RPE Cell Line

Abstract

Abstract: : Purpose: RPE cells typically have limited division potential in culture and senesce around passage 30. This limits the types of experiments that can be performed on cultured RPE cells. Although a human RPE cell line that has been transformed through transfection with DH5-HPV-16 is commercially available, it is possible that viral transfection alters the cells' phenotype. We report here on the characterization of a spontaneously transformed bovine RPE cell line. Methods: Primary RPE cells were obtained from fresh calf eyes (Brown Packaging, S. Holland, IL). The cells were plated in multi-well tissue culture dishes and grown at 37^oC, 2% CO₂ until confluent. The cultures were re-fed every three days with DMEM + 10% FBS and antibiotics. The cultures were passaged by trypsinization. Chromosome analysis and autoradiography have been used to characterize the cell line. Results: We have isolated a bovine RPE cell line that appears to be spontaneously transformed. The cell line has undergone over 100 population doublings since it was first isolated but still exhibits contact inhibition. Autoradiography experiments indicate that the rate of growth slows down with confluency suggesting that the cells are partially transformed (3T3-like). The cells can be re-pigmented with ocular melanin and it is possible to perform colony forming experiments with them. There is also a variation in chromosome number within the cell population which is indicative of spontaneous transformation. Conclusions: The isolated RPE cell line appears to be partially transformed. It exhibits some characteristics of non-transformed cells such as contact inhibition but the cells have not senesced after over 100 population doublings. This cell line should provide an excellent model for the study of RPE cells in culture and will allow more extensive experiments to be performed.

Keywords: retinal pigment epithelium • retinal degenerations: cell biology • retina