Influence of Donor Age on Establishing Well-Differentiated Clonal Cultures from Embryonic Chicken Retinal Pigmented Epithelium. (pigment cell/eye/embryo/cell culture/development)

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Abstract
Retinal pigmented epithelia (RPE) isolated from chicken embryos of various developmental stages were dissociated into single cells, and their ability to re-express defferentiated characteristics in clonal culture was investigated. The lightly pigmented, columnar cells isolated from stage 25 to 29 embryos dissociated more easily than the heavily pigmented, cuboidal cells from embryos of stages 30 to 34. The yield of RPE cells per embryo increased with donor age, paralleling the growth of the epithelium in vivo. However, the potential for these cells to attach, to proliferate, and to form typical, well-differentiated RPE colonies declined with donor age. Cells from stage 25 embryos developed exclusively into large, typical epithelial colonies which expressed all stages of differentiation from flat, unpigmented cells at the margin to cuboidal, pigmented cells in the centre. At the other end of the spectrum, cells from stage 34 embryos frequently formed small, atypical colonies of unpigmented cells, in addition to typical but relatively small colonies. The plating efficiency (calculated on the basis of pigmented colonies formed within 3 weeks) dropped from more than 2% at stage 25 to 0.01% at stage 34.