Gonzalez Cordero, A; (2013) Retinal repair using ES cell-derived photoreceptor precursors. Doctoral thesis (PhD), UCL (University College London).

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Abstract

Cell transplantation is a promising strategy for treating blindness caused by the loss of photoreceptors. Recent work by other members of the lab has demonstrated the restoration of visual function following the transplantation of photoreceptor precursors obtained from early post-natal retinas into adult GNAT1-deficient mice. This thesis has sought to investigate the use of Embryonic Stem (ES) cells as a source of photoreceptors. The studies presented here aim to (1) characterize ES derived retinal progenitors development, (2) determine the stage of development of ES cell- derived rod photoreceptor precursors by comparing it to normal development and to (3) demonstrate the capability of integration and maturation of ES cell derived photoreceptor precursors following transplantation to the adult degenerate retina. To provide a comprehensive understanding of the ES differentiation system in 2D and 3D cultures, we analysed the differentiation of ES cells into retinal progenitor cells (RPCs) and retinal cell types. Photoreceptor genesis was characterized at the transcriptional and protein levels not only to establish if these cultures followed normal development but also to find the appropriated time point for transplantation. Recombinant AAV2/9 viral vector was used to label ES cell-derived retinal cell types not only to enrich for a pure population of retinal cells but also to allow their identification after subretinal transplantation in to models of retinal degeneration. These studies demonstrated that retinal development in vitro resembles normal development, following a conserved order of birth in both differentiation cultures. ES cell-derived photoreceptors are comparable to early post-natal stages of development. Following transplantation, 2D generated retinal cell types did not integrate into the host adult retina, whereas ES cell-derived photoreceptors generated from 3D cultures were able to integrate into the outer nuclear layer and demonstrated mature photoreceptor morphology. Therefore, ES cells can be used as a source of integration competent photoreceptor precursors.

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