Retinitis pigmentosa (RP) is a hereditary disease that causes the gradual degeneration of the photoreceptor cells in the retina, starting with the rods, leading to a progressive loss of vision over time. RP is the most prevalent form of inherited retinal dystrophy, affecting over 1.5 million individuals worldwide and significantly impacting both patients and society. RP is a primary cause of visual disability and blindness in people under 60 years old, and common symptoms include nyctalopia and gradual peripheral vision loss. If left untreated, RP can ultimately lead to complete blindness.
Cell therapy is a potential method for vision restoration that can be achieved through two forms: (1) replacing dysfunctional cells with effective stem cells and (2) restoring the dysfunctional cells by releasing trophic factors. Successful integration and the formation of new synaptic connections with the host are essential for transplanted cells to be functional .
Embryonic stem cells (ESCs) are pluripotent stem cells that can self-renew and develop into the three primary germ cells. However, ESCs can be rejected by the immune system. Induced pluripotent stem cells (iPSCs) provide an alternative source that can differentiate into any somatic cell and are generated from a somatic cell line without using human embryos. Dysfunction of the retinal pigment epithelium causes significant vision loss by affecting photoreceptors. Replacing damaged retinal pigment epithelium and photoreceptors with healthy pluripotent stem cells can delay disease progression and potentially restore vision loss .
In 2004, rat models demonstrated the potential of iPSC implantation to improve vision. In 2021, Surendran et al. investigated the effects of hiPSC-derived retinal cells in mice, observing improved photoreceptor progenitor and retinal pigment epithelium functions,increased pigmentation and ciliation, tight junction protein expression, and growth factor secretion. Retinal organoid transplantation in rats also showed significant visual function improvement and integration into the host. Recent studies have demonstrated the efficacy of hiPSC-RPE cells in slowing the progression of retinal degeneration in a swine model. In addition, combining retinal pigment epithelium (RPE) and retinal sheets in RSC rats can fully replace the degenerated retina, making it a potential therapy for advanced retinal degeneration.
The use of iPSCs has several advantages over using human embryos, such as ethical concerns and the ability to match patients based on blood type. However, some drawbacks should be considered. iPSCs may retain gene expression from the original cells, which can affect properties like senescence and proliferation (epigenetic memory). Furthermore, iPSCs' ability to proliferate indefinitely raises safety concerns, including the formation of teratomas.
Bone marrow stem cells (BMSCs) are a type of multipotent stem cell found in bone marrow, including mesenchymal stem cells (MSCs) and hematopoietic stem cells (CD34+ cells). BMSCs are mainly studied for their potential in treating retinitis pigmentosa (RP) due to their ability to release anti-angiogenic and neurotrophic factors, as well as immunomodulatory proteins like insulin-like growth factor-1 (IGF-1), MHC class II antigens, and Th2-related cytokines. While the ability of BMSCs to fully differentiate into photoreceptors is still under investigation, preclinical studies have shown promising results in tracking PKH26-dyed cells and observing their integration and counteraction of inflammation in mouse models. Overall, BMSCs have the advantage of being easily extracted and manipulated and have the ability to migrate towards lesion sites and trans-differentiate.
Stem cells have the potential to provide trophic support for photoreceptors. Early clinical trials using encapsulated RPE cells producing ciliary neurotrophic factors have shown photoreceptor protection in patients with RD. In vivo studies on mice have shown that transplanting MSCs into the vitreous can improve the survival and preservation of photoreceptors, delaying the progression of RP. Transplanting both fetal RPE cells and MSCs has significantly improved ERG results and increased the survival rate of transplanted cells. Bioengineering of cells is a promising strategy to increase the number of photoreceptor cells and promote the angiogenesis needed for repair and regeneration. Conjunctiva mesenchymal stem cells have been induced into photoreceptor-like cells in fibrin gel to form 3D scaffolds. These studies indicate that coculture transplantation is more beneficial compared to single-cell transplantation.
Retinal progenitor cells (RPCs) are multipotent stem cells derived from human fetuses between 16 to 20 weeks of gestation, and can be induced to differentiate into photoreceptors and other neuronal cells of the retina in vitro . Transplantation of RPCs has been shown in preclinical studies to improve visual function and form functional synapses with host cells, indicating the potential to stabilize or reverse progressive vision loss . In particular, C-Kit+ RPCs derived from organoids have shown exceptional results in forming functional synapses with host cells in mice . Additionally, RPCs secrete trophic factors that can promote retinal survival and photoreceptor replacement .
Stem cell therapy has shown promising potential for regenerating the retina in preclinical studies, with the ability to differentiate retinal progenitor cells into photoreceptors and integrate transplanted cells into the degenerating retina. The formation of functional synapses between host and transplanted cells also suggests the potential for stabilizing or reversing progressive vision loss. While clinical trials are yet to be conducted, these findings hold great promise for the future of stem cell therapy.
Retinal gene therapies have limitations that prevent them from compensating for all cases of visual function loss, such as patients without genetic diagnoses and those with advanced diseases and no remaining photoreceptors. Optogenetics is a more universal therapy that can be used in such cases, and it involves converting non-light-sensitive retinal cells into artificial photoreceptors by introducing light-sensitive proteins, or opsins, into the cells. There are different types of opsins that can be used depending on the target cell and method employed. Depolarized opsins can mimic an "on" response, while hyperpolarized opsins can be used on dormant cells. Opsin genes used for optogenetic vision restoration can be categorized into two superfamilies: microbial opsins (Type 1) and animal opsins (Type 2). These opsins differ in their light sensitivity, function, and utility for vision restoration. Type 1 opsins directly affect ion channels or pumps upon light absorption, while Type 2 opsins cause intracellular G-protein-coupled receptor signaling cascades that indirectly affect ion channels.
Garita-Hernandez et al. conducted a study to evaluate the efficiency and toxicity of different microbial opsins in human retinal organoids derived from hiPSCs . Various depolarizing and hyperpolarizing microbial opsins, including CatCh, ChrimsonR, ReaChR, eNpHR 3.0, and Jaws, were tested. eNpHR 3.0, ReaChR, and Jaws exhibited the highest membrane localization, while CatChR and ChrimsonR showed lower membrane localization due to protein accumulation in the cytosol at high doses, leading to UPR . These findings suggested that hiPSC-derived retinal organoids are useful for predicting optogenetic protein expression patterns and their effects on human retinal contexts. Recent advances in optogenetic therapy using HEK293 triple-transfected cells and/or hiPSC-derived photoreceptors equipped with microbial opsins have shown promising results in improving vision and restoring cells in Cpfl1/Rho−/− mice and wild-type mice (C57BL/6J) .
Ganglion cells have been the focus of several large animal studies investigating vision restoration. Modified mammalian ionotropic glutamate receptor (LiGluR) was delivered to the ganglion cells of Rcd1 dogs through intravitreal injection, resulting in restored vision in visible light. The delivery of maleimide-azobenzene-glutamate0 modified LiGluR resulted in acquired properties of light-gated channels and showed evidence of derived light responses. In a similar approach, optogenetic therapy was delivered to target the outer retina in Rdc1 dogs and C57BL/6 mice, demonstrating partial improvement in vision restoration through Y-maze performance and electroretinography.
Promoters play a crucial role in optogenetics. In macaque retinas, the combination of the ubiquitous cytomegalovirus promoter with an AAV2 vector increased upregulation of ChR-Ca2+-permeable ChR, improving light sensitivity. The gamma-synuclein gene promoter region was found to increase CatCh expression in ganglionic cells. The optimization of ChRs' light sensitivity through two CoChR mutants (CoChR-L112C and CoChR-H94E/L112C/K264T) delivered with an AAV2 vector restored light sensitivity in TKO mice. Chloromonas oogama, a variant of green algae, has been shown to improve light sensitivity in blind mice and restore vision under ambient light conditions.
Optogenetic therapy for vision restoration has potential safety concerns, such as phototoxicity induced by continuous light exposure and UV light. However, a study on mice transduced with optogenetic gene mVChR1 (using an AAV vector) and exposed to continuous light for one week showed no reduction in visually evoked potentials (VEPs) amplitudes in RGCs and an absence of phototoxicity. Mutations in opsins were also explored, and the third extracellular loop in mVChR1 was found to be implicated in sodium ion selectivity, and transmembrane 6 and C-Terminal regions have a role in some channel kinetics. G-protein stimulation has also shown to restore light sensitivity. Different optogenetic variants and promoters have been studied, providing potential treatment options for patients based on their specific needs. Further testing is needed to determine amino acid sequences that determine wavelength sensitivity.
In brief, optogenetics has been shown to partially restore vision in animal models of RP, but its safety and efficacy in humans remain to be fully understood as it is still in the preclinical stage. Furthermore, using various variants of opsins and promoters may improve the outcome of optogenetics, providing more options for patients with RP.