Abstract

This study was designed to determine whether adult mouse induced pluripotent stem cells (iPSCs), could be used to produce retinal precursors and subsequently photoreceptor cells for retinal transplantation to restore retinal function in degenerative hosts. iPSCs were generated using adult dsRed mouse dermal fibroblasts via retroviral induction of the transcription factors Oct4, Sox2, KLF4 and c-Myc. As with normal mouse ES cells, adult dsRed iPSCs expressed the pluripotency genes SSEA1, Oct4, Sox2, KLF4, c-Myc and Nanog. Following transplantation into the eye of immune-compromised retinal degenerative mice these cells proceeded to form teratomas containing tissue comprising all three germ layers. At 33 days post-differentiation a large proportion of the cells expressed the retinal progenitor cell marker Pax6 and went on to express the photoreceptor markers, CRX, recoverin, and rhodopsin. When tested using calcium imaging these cells were shown to exhibit characteristics of normal retinal physiology, responding to delivery of neurotransmitters. Following subretinal transplantation into degenerative hosts differentiated iPSCs took up residence in the retinal outer nuclear layer and gave rise to increased electro retinal function as determined by ERG and functional anatomy. As such, adult fibroblast-derived iPSCs provide a viable source for the production of retinal precursors to be used for transplantation and treatment of retinal degenerative disease.

Highlights

  • Retinal degenerative diseases such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD) are currently the leading cause of incurable blindness in the western world [1,2,3]

  • One significant achievement came in 2004 when a subset of transplanted retinal progenitor cells was shown to develop into a variety of mature retinal neurons, including retinal ganglion and photoreceptor cells [4]

  • Numerous studies reporting varying degrees of success have utilized an assortment of different cell types ranging from the fate-restricted photoreceptor precursor [5] to the pluripotent embryonic stem (ES) cell [6,7,8,9]

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Summary

Introduction

Retinal degenerative diseases such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD) are currently the leading cause of incurable blindness in the western world [1,2,3]. The originally published protocols remain the most effective and efficient means of inducing pluripotency in adult dermal fibroblasts, a variety of new protocols with variations in cell type, delivery methods and reprogramming factors have been developed [11,12,13,14,15,16,17,18,19]. Rod and cone photoreceptor cell repopulation, synapse formation and cellular integration associated with restoration of electrophysiological and anatomical correlates of retinal function were observed These findings establish a proof-of-principle for applications of autologous iPSCs for the safe and effective treatment of retinal degenerative diseases

Results and Discussion
Materials and Methods
Methods

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