As a general principle, patients who could directly benefit from cell-based therapies with retinal degenerative disease such as RP and AMD will require replacement of lost photoreceptor cells, RPE, and possibly choriocapillaris [160]. Korte et al. have shown the choriocapillaris can regenerate if areas of absent RPE can be repopulated with new RPE. Thus the clinical need here is to promote the differentiation of stem cells into photoreceptors (rods and cones) and RPE.

In dry AMD, loss of vision arises from loss of RPE and photoreceptors with secondary atrophy of choriocapillaris. A potential treatment for AMD and inherited disease that affect the RPE and photoreceptors would be cell replacement therapy, but one significant hindrance to the clinical use of cell transplantation for treatment of retinal degenerations is the availability of a source of replacement cells. Although RPE derived from prenatal and postnatal tissue has been isolated and induced to grow in vitro, such sources are limited and vary in terms of quality and expansion capacity [141, 161–163]. Moreover, it has been demonstrated that postmitotic photoreceptor precursor cells can be derived from tissue of the early postnatal mouse retina (P1–P5) [164, 165]. However, equivalent retinal cells in humans would have to be derived from second-trimester fetuses. While these studies provide solid evidence that transplantation strategies show great potential, an approach such as this would have ethical implications as well as the problem of a limited reservoir of donor cells [165].

Stem cells are an excellent source of cells for replacement therapy given the limited reservoir of donor cells for RPE replacement strategies, lack of regeneration of photoreceptors, and variation in success of autografts. Stem cells have been isolated from a variety of sources including embryo and adult eye [164, 166]. Human embryonic stem cells (hESC) are being investigated as a potential source of photoreceptors and RPE and are promising candidates for therapeutic use.

As mentioned, the strategy in cell replacement therapy using stem cells is to differentiate these cells into photoreceptors or RPE. The extracellular environment plays a critical role in the differentiation of stem cells into the target cell type and extracellular matrix can differentiate cells into the adjacent cell layer. For example, hESCs cultured on a monolayer of cells derived from mouse calverium can be induced to a neural fate and express neural progenitor markers such as paired boxed protein (PAX)-6, neurofilament, and glial fibrillary acidic protein (GFAP) (Fig. 1.6) [167]. Similarly, hESCs cultured on a monolayer of RPE (ARPE19) cells can be induced to express neural retinal markers such as vimentin, neurofilament, and cone–rod homeobox (CRX), which is essential in early photoreceptor development (Fig. 1.7) [167]. Varying extracellular matrix environs such as laminin, matrigel, and/or vitronectin and fibronectin can also induce embryonic stem cells toward a neural progenitor or RPE fate. Embryonic stem cells cultured on laminin, vitronectin, and fibronectin can be induced to express neural progenitor markers such as neurofilament and neural retina-specific leucine zipper (NRL), an intrinsic regulator of photoreceptor development (Fig. 1.8). These cells can also be induced to express RPE markers such as tight junction protein ZO-1 and bestrophin when cultured on matrigel (Fig. 1.9). Stem cells grown on human Bruch’s membrane have also been induced to differentiate to RPE (Fig. 1.10) [167].

Current clinical trials are underway to evaluate the potential of stem cell therapy in humans. Advanced Cell Technology, Inc. initiated a phase 1 clinical trial in humans in 2011 for the treatment of retinal degenerative disorders [141]. This is the first FDA approved trial for the treatment of macular degeneration using RPE derived from human embryonic stem cells. RPE cells were derived from a single donor human embryonic stem cell line. The preliminary report details the phase 1 trial being conducted to test the safety and tolerability of hESC-RPE in patients with advanced-stage Stargardt’s macular dystrophy and dry age-related macular degeneration. Briefly, a human embryonic stem cell (MA09) line was first used to generate hESC-derived RPE, which were then characterized and tested for pathogen contamination. After pars plana vitrectomy, submacular injections of 50,000 cells were used to treat the patient. Patients were immune suppressed with low-dose tacrolimus and mycophenolate mofetil 1 week before surgery and continued for 6 weeks postsurgery. After 6 weeks patients discontinued tacromilus and continued a mycophenolate regimen for 6 more weeks. Both patients tolerated the injection without signs of postoperative inflammation, rejection, or tumorigenicity at time of the report (4 months follow-up). In the patient with Stargardt’s macular dystrophy, transplanted cells attached to Bruch’s membrane and persisted throughout the observation period; there was possible visual improvement in the injected eye as shown by visual acuity and Goldmann visual field test. In the AMD patient no clinically detectable sign of successful transplantation was observed, although the patient did not comply with the immunosuppressive drug regimen [168]. Interestingly, there was mild visual improvement in both eyes of the patient with AMD as shown by visual acuity and Goldmann visual field test. It is unclear whether these visual improvements are due directly to the cell transplant or a secondary cause such as the immunosuppressive drugs or a placebo effect [141]. Nevertheless, these important studies demonstrate that stem cells can be transplanted into the subretinal space in humans without abnormal proliferation, teratoma formation, graft rejection, or other untoward pathological reaction or safety signal.

These short-term results in only two patients, are preliminary but provide valuable proof-of-concept evidence for future treatment of macular degeneration and Stargardt’s disease in humans. Long-term follow-up in a larger cohort of patients is needed to draw more meaningful conclusions from this trial [168]. Further trials are also needed to determine the optimal number of transplanted cells, immunosuppression regimens, and disease stage for transplantation [168].

The recent development of induced pluripotent stem (iPS) cells holds great promise as a potential reservoir of cells for the treatment of age-related macular degeneration and other disorders. Induced pluripotent stem cells were initially generated in 2006 by the Yamanaka group and the technology has had dramatic implications both from an ethical and scientific standpoint [169]. The technology is a significant advancement over prior technology, as it allows researchers to generate pluripotent cells for potential therapeutic use without the controversial use of embryos. These cells are generated by reprogramming adult somatic cells using transcriptional regulators such as SOX2, OCT3/4, and Klf4 [169, 170]. These cells are then reprogrammed with similar potential as embryonic stem cells and are capable of differentiating into three germ layer cell types (mesoderm, ectoderm, and endoderm) [169, 171, 172]. These iPS cells hold great promise for the generation of RPE and photoreceptors for cell replacement therapy and create a new paradigm as a novel reservoir. Tsang et al. have generated RPE-like cells from human iPS (Fig. 1.11). Several other groups have used iPS technology to generate photoreceptors and then transplant these cells into animal models of retinal degeneration [173–175].

Induced pluripotent stem cells can also provide a platform to study disease through the use of patient-specific iPS cells. Through the generation of iPS cells from patients with specific diseases, models can be developed to express particular disease phenotypes which can then be used to understand pathophysiology of disease and determine the efficacy of therapeutic interventions [176]. These models can also be developed to help understand human inherited diseases given their clinical and genetic heterogeneity [177]. Cells derived from a particular patient can be used as a biological tool for drug discovery and toxicity testing of therapeutic agents, providing a new paradigm for personalized medicine [176].

Utilizing iPS cells as a tool for cell replacement therapy could also reduce the possibility of immune rejection given their autologous nature. Use of patient-specific iPS-derived RPE, generated from somatic cells of the potential transplant recipient with geographic atrophy, has one major and important theoretic advantage over other potential cell sources, namely, the avoidance of graft rejection. This is an important advantage, since long-term systemic immune suppression is poorly tolerated in elderly patients. Although the presence of anterior chamber-associated immune deviation (ACAID) confers some immune privilege in the subretinal space, allogeneic RPE will undergo graft rejection after subretinal transplantation unless immune suppression is used [178, 179]. Use of patient-specific iPS may circumvent graft rejection, which is one of the major challenges to ensuring graft survival in the subretinal space.

Challenges remain to the successful use of iPS cells. Induced pluripotent cells derived from the affected patient contain the predisposing mutation that caused the disease. This can provide a unique disease model but the mutation may also impede the function of the transplanted cells. These stem cells may have to first be repaired by targeted gene therapy or other techniques prior to transplantation. Additional work is also needed to translate the advances of iPS cells into clinical trials to assess safety and efficacy. Better understanding of iPS cell technology and refining the methodology of their generation will have a significant impact on retinal degenerations and regenerative medicine.