The future of regenerative medicine looks bright. We are gaining new understanding of the potential of stem cells for tissue repair in a wide range of biological systems. Recent and remarkable breakthroughs include the discovery that somatic cells can be reprogrammed to a pluripotent state via the expression of a cocktail of transcription factors Citation[1,2]. The generation of these induced pluripotent stem (iPS) cells provides a previously unimaginable regenerative potential. Novel research methods, for example, to differentiate human embryonic stem (hES) cells into specific types of cells that are lost or abnormal in disease states, are creating new therapeutic options Citation[3]. Meanwhile, innovation in translational techniques such as the delivery of newly synthesized cells to diseased tissue and the bioengineering of complex tissues in vitro are being accelerated by the rapid progress in stem cell biology research and have regularly received media attention.
The visual system is a prime target for stem cell therapy for a number of compelling reasons. The scale of the global challenge presented by untreatable blindness involving the death of retinal neurons, either the light-sensitive photoreceptors or the retinal ganglion cell projection neurons, provides a strong impetus. More than 3.2 million people are currently blind from age-related macular degeneration, and this number is set to rise in aging populations Citation[4]. A total of 1.7 million people are blind from diabetic retinopathy and this figure is also rising in developing countries with changing economic and lifestyle factors Citation[4]. One in 3000 people have an inherited retinal disease caused by mutations in any one of more than 200 different genes Citation[101]. Conversely, loss of retinal ganglion cells in glaucoma causes blindness in approximately 4.5 million people Citation[4]. The eye is a highly tractable system for the delivery of new cell populations through existing advanced surgical techniques, and this is relatively straightforward compared with delivery to less accessible regions of the CNS. The developmental biology and genetics of the eye are intensively studied, and the research has provided knowledge of a repertoire of intrinsic and extrinsic factors that regulate the generation of different retinal cell types that is now being deployed in an effort to regenerate retinal tissue and generate new cells in vitro. The first gene therapy trials for an inherited retinal disease were conducted in 2008 Citation[5–7] and have set the scene for pioneering new therapies for retinal disease. The success of gene therapy relies on the delivery of new functional genes to cells that lack such genes, and is therefore entirely dependent upon cell survival. In cases where the degenerative process has already culminated in cell death or in those conditions that are not amenable to gene therapy approaches, stem cell therapies offer a complementary approach.
There are two related yet distinct goals of stem cell therapy for retinal disease. The most challenging and sought-after goal is to replace the retinal neurons that have died with new functional neurons that are able to repopulate the diseased retina and that are functionally indistinguishable from normal retinal cells. This approach offers the prospect of restoration of sight. The alternative goal is to transplant cells that are able to preserve the function of the diseased tissue and prevent cell death, thereby prolonging visual function. The former approach also shares the latter’s goal as the new functional neurons also face the challenge of long-term survival in a potentially hostile degenerating environment.
In both approaches, a central requirement is to identify the type of cell that will be deployed to repair the retina. Of considerable interest is the idea that endogenous cells in the eye itself could be recruited to regenerate diseased tissue. Such cells would have the significant advantage of avoiding the rejection problems associated with allotransplantation. In non-mammalian lower vertebrates, neurogenesis generating new retinal neurons continues throughout life. Moreover, regeneration following injury is possible and draws on different stem cell populations in different species. However, thus far it has not proven possible to mimic these regenerative capacities via stimulation of cells in the mammalian retina, which cease retinal neurogenesis before birth in human development. Attention has been focused on stem cells of the ciliary epithelium that can be expanded in vitro to increase cell number, but their ability to generate new differentiated retinal neurons appears limited both in vivo and in vitroCitation[8–11]. The alternative method to regenerating tissue from cells within the eye itself is instead to transplant new cells into the diseased environment. This goal has been historically pursued using retinal tissue from external sources from either fetal or adult donors and suffers from the limited availability of tissue. One of the compelling reasons for the ascendancy of the use of stem cells is the prospect of generating unlimited quantities of new cells from immortal stem cell lines. In addition to the investigation of ciliary epithelium cells, a number of other putative stem cell sources have been studied as potential sources of new retinal cells. Over the last 3 years, it has become clear that the best source identified to date is the pluripotent embryonic stem cell derived from the blastocyst of the early-stage embryo. Several recent studies have produced very convincing demonstrations of the generation of photoreceptors and other retinal neurons in vitro from human, monkey and mouse ES cells using related protocols Citation[2,12,13]. Similar differentiation has been achieved using iPS cells generated by reprogramming skin fibroblasts or other somatic cells, although the iPS field is in its infancy and our understanding of the epigenetic mechanisms underlying reprogramming and its efficacy remain to be elucidated.
With this new-found ability to generate new retinal cells in vitro that could potentially replace those lost through disease, the importance of understanding whether such cells could be persuaded to integrate with the retinal circuitry of a host retina is paramount if cell-replacement therapy using stem cell sources could ever be achieved. It is in answering this question that our own work has made a significant contribution. By transplanting cells isolated from the developing retina at a stage when rod photoreceptors are normally newborn and exist as immature cells, we were able to show that the adult non-neurogenic retinal environment is able to support the maturation of new photoreceptors. Furthermore, these newly matured cells were able to connect to the inner retinal neurons and confer increased sensitivity to light in the rod range Citation[14]. In addition, the new photoreceptors show long-term survival and the number of new rod cells that integrate into the host outer nuclear layer can be enhanced by modulation of the outer limiting membrane Citation[15,16]. Importantly, these results were only achieved using photoreceptor precursor cells that are specified to differentiate into photoreceptors but not yet exhibiting mature morphological features of photoreceptors such as outer segments and synapses at the time of transplantation. The same result was not possible using proliferating stem cells as these failed to differentiate. More recently, we showed that it is possible to transplant cones as well as rods also using immature precursor cells Citation[17]. These discoveries define a strategy for photoreceptor cell-replacement therapy and have overcome a number of obstacles. They show that cell replacement is feasible provided the correct stage cell is transplanted. As this cell type, which we call a photoreceptor precursor, is postmitotic, it avoids the potential hazards associated with tumor formation owing to unregulated proliferation of transplanted undifferentiated stem cells.
In our opinion, the best current strategy for photoreceptor cell replacement for retinal disease is to generate photoreceptor precursors from hES cell lines, and to transplant these immature cells into the subretinal space from where they can differentiate and integrate into the host retinal circuitry to generate new functional photoreceptors. hES cell lines are derived from human eggs that are fertilized in vitro and are immortal. Already, there have been transplantations of hES-derived cells in mouse models, which show promising results Citation[18]. The challenge now is to ensure that the hES-derived cells are as effective as cells from the developing retina by selecting and transplanting only photoreceptor precursor cells at the correct developmental stage.
The first clinical trial of hES-derived cell transplants into patients with Stargardt’s macular dystrophy was approved in 2010 in the USA, and a similar study is planned in the UK. These trials plan to transplant the retinal support cells, the retinal pigmented epithelial (RPE) cells into the eye in order to replace degenerated RPE cells and support photoreceptor survival Citation[19,20], rather than transplanting new photoreceptors. Critically, they will test the safety and tolerance of the ES-derived RPE cells following subretinal injection in the eye. If these trials show a risk-free outcome, they will set the stage for further clinical trials exploring the use of ES-derived cells in retinal disease. Although there are many obstacles to overcome, especially in determining which diseases and stages of disease are likely to benefit from stem cell therapies, and how effective and for what period of time such treatment could give improved outcomes, there is much room for optimism. The field is entering an unprecedented period of discovery and innovation for the development of new stem cell therapies for blinding conditions that were once considered to be untreatable and incurable.
Financial & competing interests disclosure
Jane C Sowden and Robin R Ali have been awarded grants from the following: MRC grants GO3000341 and G0901550; NIHR Biomedical Research Centre for Ophthalmology at Moorfields Eye Hospital and UCL Institute of Ophthalmology; Great Ormond Street Hospital Children’s Charity. The authors have no other relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript apart from those disclosed.
No writing assistance was utilized in the production of this manuscript.
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Website
- RetNetTM. Retinal Information Network www.sph.uth.tmc.edu/retnet