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Arnold Szabo, Akos Kusnyerik, Ferenc Kilin, Akos Lukats, Janos Nemeth, Zoltan Zsolt Nagy, Agoston Szel; Long-term organotypic co-culture model of the adult human retina, RPE and choroid. Invest. Ophthalmol. Vis. Sci. 2018;59(9):4021.
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© ARVO (1962-2015); The Authors (2016-present)
Corneal transplantation replaces diseased tissue with healthy tissue from an organ donor. While the surgery to harvest transplantable corneas from recently deceased donors removes the whole eye, the rest of the tissue is typically disposed of. We developed a culture technique which maintains retinas from those eyes in excellent condition for more than three months, enabling research on human retinal tissue that was previously impossible.
Human eyes were collected through scheduled multi-organ donations. Following enucleation and dissection of the eye ball, organotypic retina cultures were prepared within 1-2 hours after the circulation arrest in the donor. The retinal pieces with or without attached retinal pigment epithelium (RPE) and choroid were cultured for up to 100 days in a specific, serum-free, chemically defined medium which has been optimized for the human retina.
The morphology of both isolated neural retina and retina-RPE-choroid co-cultures were astonishingly well preserved with low inter-sample variability. Every major cell type survived and all retinal layers were maintained even after twelve weeks. Outer segments could be detected in high numbers in both cultures, but the quality and the density of outer segments were superior in retina-RPE-choroid co-cultures. Cones did not undergo severe apoptosis and a mean density of 5000-5500 cones/mm2 were measured even in long-term cultures. Subpopulations of bipolar, horizontal and amacrine cells showed close to normal morphology. The stratification of the inner plexiform layer remained recognizable. Only the number of surviving ganglion cells showed a significant decrease in long-term cultures, but ganglion cells were still present even after 84 days. Synaptic structures showed close to normal morphology on samples stained against synapthophysin, while prominent telodendria on cone pedicles indicated intact gap junctions.
The adult human retina can be maintained in an appropriate culture system for at least three months. By long-term culturing, both acute and chronic effects of pharmacological compounds could be tested directly on human tissue in a cost- and time-effective manner. Further, the long culture time allows the administration of viral vectors and opens new strategies for developing and testing gene therapeutic approaches and can help to reduce the use of animals both in academic and industrial research.
This is an abstract that was submitted for the 2018 ARVO Annual Meeting, held in Honolulu, Hawaii, April 29 - May 3, 2018.
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