Goh, Debbie; (2017) Assessing visual function following transplantation of mouse ESC-derived rod photoreceptor precursors. Doctoral thesis (Ph.D), UCL (University College London).

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Abstract

Photoreceptor replacement therapy is a promising strategy for treating retinal degenerative diseases leading to blindness. Previous work by our lab has demonstrated proof-of-concept that mouse embryonic stem cells (ESCs) can be differentiated, in a 3D culture system, into post-mitotic rod photoreceptor precursors for transplantation. However, rescue of visual function following transplantation of ESC-derived rod precursors has yet to be demonstrated. This project therefore sought to optimise the transplantation of mouse ESC-derived rod photoreceptor precursors into mouse models of retinal disease, define the optimal developmental stage for transplantation in an endogenous photoreceptor reporter ESC line, and thereafter, assess the extent to which visual function is restored in recipient animals post-transplantation. We assessed different aspects of optimising the transplantation protocol, such as dissociation method, AAV2 virus pseudotype used for labelling photoreceptors, and host immune suppression, and also characterised photoreceptor differentiation in a Crx.GFP ESC line. Additionally, we developed a multi-electrode array (MEA) set up and stimulus protocol that allowed us to detect visual responses from retinal explants. These studies demonstrated the generation of large numbers of healthy, developmentally-homogenous mouse ESC-derived rod photoreceptors for transplantation, and resulted in a significantly increased number of donor-reporter labelled cells observed in the host outer nuclear layer (ONL) following transplantation. Most importantly, using the MEA, we were able to demonstrate for the first time that mouse ESC-derived photoreceptor precursors were capable of transmitting light-evoked responses following transplantation into a mouse model of severe retinal degeneration. Taken together, these are encouraging first steps towards the use of stem cells for photoreceptor replacement therapy in retinal degenerative diseases.

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