title: Enhanced functional integration of human photoreceptor precursors into human and rodent retina in an ex vivo retinal explant model system.
abstract: Retinal disease is the major cause of irreversible blindness in developed countries. Transplantation of photoreceptor precursor cells (PPCs) derived from human embryonic stem cells (hESCs) is a promising and widely applicable approach for the treatment of these blinding conditions. Previously, it has been shown that after transplantation into the degenerating retina, the percentage of PPCs that undergo functional integration is low. The factors that inhibit PPC engraftment remain largely unknown, in part, because so many adverse factors could be at play during in vivo experiments. To advance our knowledge in overcoming potential adverse effects and optimize PPC transplantation, we have developed a novel ex vivo system. Harvested neural retina was placed directly on top of cultured retinal pigment epithelial (RPE) cells from a number of different sources. To mimic PPC transplantation into the subretinal space, hESC-derived PPCs were inserted between the retinal explant and underlying RPE. Explants cocultured with hESC-derived RPE maintained normal gross morphology and viability for up to 2 weeks, whereas the explants cultured on ARPE19 and RPE-J failed by 7 days. Furthermore, the proportion of PPCs expressing ribbon synapse-specific proteins BASSOON and RIBEYE was significantly higher when cocultured with hESC-derived RPE (20% and 10%, respectively), than when cocultured with ARPE19 (only 6% and 2%, respectively). In the presence of the synaptogenic factor thrombospondin-1 (TSP-1), the proportion of BASSOON-positive and RIBEYE-positive PPCs cocultured with hESC-derived RPE increased to ∼30% and 15%, respectively. These data demonstrate the utility of an ex vivo model system to define factors, such as TSP-1, which could influence integration efficiency in future in vivo experiments in models of retinal degeneration.
authors.affiliation: Department of Ophthalmology and Visual Sciences, Faculty of Medicine, University of British Columbia, Vancouver, British Columbia, Canada.
authors.forename: Anat
authors.initials: A
authors.lastname: Yanai
authors.affiliation: Department of Ophthalmology and Visual Sciences, Faculty of Medicine, University of British Columbia, Vancouver, British Columbia, Canada.
authors.forename: Christopher R J
authors.initials: CR
authors.lastname: Laver
authors.affiliation: Department of Ophthalmology and Visual Sciences, Faculty of Medicine, University of British Columbia, Vancouver, British Columbia, Canada.
authors.forename: Cheryl Y
authors.initials: CY
authors.lastname: Gregory-Evans
authors.affiliation: Department of Ophthalmology and Visual Sciences, Faculty of Medicine, University of British Columbia, Vancouver, British Columbia, Canada.
authors.forename: Ran R
authors.initials: RR
authors.lastname: Liu
authors.affiliation: Department of Ophthalmology and Visual Sciences, Faculty of Medicine, University of British Columbia, Vancouver, British Columbia, Canada.
authors.forename: Kevin
authors.initials: K
authors.lastname: Gregory-Evans
mesh_terms: D000429:Alcohol Oxidoreductases / Q000096:biosynthesis / Q000235:genetics; D000818:Animals; D002454:Cell Differentiation; D002460:Cell Line; D017690:Cell Transplantation; D056970:Co-Repressor Proteins; D018920:Coculture Techniques; D004847:Epithelial Cells / Q000166:cytology; D000066449:Human Embryonic Stem Cells / Q000166:cytology*; D006801:Humans; D051379:Mice; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000235:genetics; D009924:Organ Culture Techniques; D020419:Photoreceptor Cells, Vertebrate / Q000166:cytology*; D051381:Rats; D012160:Retina / Q000166:cytology*; D055213:Retinal Pigment Epithelium / Q000166:cytology; D019700:Thrombospondin 1 / Q000378:metabolism*
chemical_list: C498440:BSN protein, human; D056970:Co-Repressor Proteins; D009419:Nerve Tissue Proteins; D019700:Thrombospondin 1; D000429:Alcohol Oxidoreductases; C421248:CTBP2 protein, human