R57 and R28: Two multipotential retinal precursor cell lines

Purpose. The purpose of this study was to obtain and characterize clonal populations of retinal cells derived from a mixed immortalized rat retinal culture. Such purified cell lines would have broad applications for in vitro studies of retinal cell growth, differentiation, and gene expression. Methods. Neuroretinae from postnatal day 6 rats were immortalized with a replication incompetent EIA-containing retrovirus as described [Seigel, G.M. 1996, In Vitro Cell. Devel. Biol. 32, in press]. Clonal cell lines R57 and R28 were obtained by three rounds of limiting dilution to ensure single cell origin. Phenotypes were analyzed immunocytochemically using a battery of retina-specific monoclonal antibodies. Results. Despite the clonal nature of these cell lines, variations in morphologies and cell staining patterns were evident. Subpopulations of R57 and R28 stained positively for photoreceptor markers IRBP, S-antigen, and recoverin. The possibility of rhodopsin expression is still under investigation. Expression of markers indicative of other retinal cell types was suggested by immunoreactivity to monoclonal antibodies with the following specificities: Ret G1 (Müller), RetB1 (bipolar, photoreceptor), 2G12 (ganglion), and 8A1 (horizontal, ganglion, amacrine). No immunoreactivity was seen for monoclonal antibodies: HPC1 (amacrine) and VC1.1 (horizontal, amacrine). Conclusions. We have successfully established two clonal cell lines which express markers consistent with at least photoreceptor, Müller, and ganglion cell phenotypes. These multipotential subpopulations suggest a precursor cell origin for both R57 and R28. We are currently conducting both in vitro, as well as in vivo studies to determine the full spectrum of their differentiation potential.