Cytoskeleton, Adhesion, and Extracellular Matrix of Fetal Human Retinal Pigmented Epithelial Cells in Culture

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Abstract
Small sheets of human retinal pigmented epithelial (RPE) cells isolated from 6- to 9-week-old fetal human eyes were used to establish RPE cells in culture. After 4-5 days in culture the RPE cells in various parts of a colony were examined by immunofluorescence and fluorescence microscopy to determine the distribution of the cytoskeletal proteins tubulin, myosin, F-actin, vimentin, and cytokeratin and the extracellular matrix proteins laminin and type IV collagen. The extracellular matrix components laminin and type IV collagen, which are abundant underneath the RPE cells in the centre of the colony, progressively decrease in amount towards the edge of the colony. Microtubules labeled by antisera to tubulin were present in the cytoplasm of RPE cells throughout the colony. F-actin and myosin were found in stress fibres in the RPE cells at the edge of the colony and in compact circumferential rings circumscribing the apical ends in RPE cells in the centre of the colony. Examination of cell-substratum adhesion patterns by interference reflection microscopy showed that focal contacts were present only in RPE cells at the edge of the colony where they were located at the ends of the stress fibres. The large focal adhesions previously observed in colonies of chick RPE cells, however, were not seen in human RPE cells. Human RPE cells throughout the colony had cytokeratin-containing intermediate filaments as well as vimentin-containing intermediate filaments, whereas only the latter were observed in chick RPE cells. These results indicate that in the general organization of the cytoskeleton and and in the distribution of the extracellular matrix components, laminin and type IV collagen, colonies of human RPE cells are similar but not identical to colonies of chick RPE cells.