Retinal pigment epithelial cells from Royal College of Surgeons dystrophicrats can take up melanin granules

Background: Many successful pigment epithelium transplantation studies invo lving pink-eyed Royal College of Surgeons (RCS) dystrophic rats showed high ly pigmented transplanted cells forming a double layer with slightly pigmen ted cells, attached to Bruch's membrane. Since it is not clear whether tran splanted pigmented cells can displace retinal pigment epithelial (RPE) host cells from Bruch's membrane, we suggested that RPE cells of RCS dystrophic rats can phagocytize melanin granules, possibly derived from perished tran splanted cells. Methods: In a series of three experiments, RPE cells of nin e pink-eyed, 21/2-month-old RCS dystrophic rats were isolated by trypsiniza tion and mechanical dissection and cultivated in Dulbecco's modified Eagles ' medium. These cells were then fed with melanin granules, isolated from bo vine RPE cells, double-trypsinized after phagocytosis and viewed by light a nd electron microscopy. We also transplanted iris pigment epithelial (IPE) cells of 20-day-old Long-Evans rats into the subretinal space of pink-eyed RCS dystrophic rats of the same age, shown in light-microscopic photography after 42 days. Results: Living RPE cells were heavily pigmented after feed ing with isolated melanin granules in all three experiments as viewed by Li ght microscopy. In addition, we identified melanin granules phagocytized by dystrophic RPE cells in electron microscopy. After transplantation of pigm ented IPE cells into the subretinal space of pink-eyed RCS dystrophic rats' eyes, a layer of slightly pigmented cells was seen on Bruch's membrane bel ow the transplanted IPE cells, shown in light microscopy. Conclusion: We ha ve shown by phagocytosis assay that dystrophic RPE cells can take up melani n granules in vitro. Our results assume that pigmented cells in transplanta tion studies, found as a monolayer, attached to Bruch's membrane, cannot au tomatically be identified as transplanted cells. Instead, the possibility o f perished transplanted cells serving as melanin donors for RPE host cells must be taken into consideration.