Ocular Stem Cells to Treat Retinal and Corneal Disorders

In this issue of Stem Cell Reports, Park et al. (2019) describe real-time in vivo visual monitoring of keratin-14+, Confetti-labeled limbal epithelial stem cells and their progeny as they contribute to central corneal wound-healing. The authors show that corneal wounds initially heal by “basal epithelial cell migration” into the wound-bed.

Maintenance of the visual function is the desired outcome of ophthalmologic therapies. The shortcomings of the current treatment options, like partial recovery, post-operation failure, rigorous post-operative care, complications, etc., which are usually encountered with the conventional treatment options has warranted newer treatment options that may eliminate the root cause of diseases and minimize the side effects. Cell therapies, a class of regenerative medicines, have emerged as cutting-edge treatment option. The corneal and retinal dystrophies during the ocular disorders are the major cause of blindness, worldwide. Corneal disorders are mainly categorized mainly into corneal epithelial, stromal, and endothelial disorders. On the other hand, glaucoma, retinitis pigmentosa, age-related macular degeneration, diabetic retinopathy, Stargardt Disease, choroideremia, Leber congenital amaurosis are then major retinal degenerative disorders. In this manuscript, we have presented a detailed overview of the development of cell-based therapies, using embryonic stem cells, bone marrow stem cells, mesenchymal stem cells, dental pulp stem cells, induced pluripotent stem cells, limbal stem cells, corneal epithelial, stromal and endothelial, embryonic stem cell-derived differentiated cells (like retinal pigment epithelium or RPE), neural progenitor cells, photoreceptor precursors, and bone marrow-derived hematopoietic stem/progenitor cells etc. The manuscript highlights their efficiency, drawbacks and the strategies that have been explored to regain visual function in the preclinical and clinical state associated with them which can be considered for their potential application in the development of treatment.

Stem cells are unspecialized cells that have been a major focus of the field of regenerative medicine, opening new frontiers and regarded as the future of medicine. The ophthalmology branch of the medical sciences was the first to directly benefit from stem cells for regenerative treatment. The success stories of regenerative medicine in ophthalmology can be attributed to its accessibility, ease of follow-up and the eye being an immune-privileged organ. Cell-based therapies using stem cells from the ciliary body, iris and sclera are still in animal experimental stages but show potential for replacing degenerated photoreceptors. Limbal, corneal and conjunctival stem cells are still limited for use only for surface reconstruction, although they might have potential beyond this. Iris pigment epithelial, ciliary body epithelial and choroidal epithelial stem cells in laboratory studies have shown some promise for retinal or neural tissue replacement. Trabecular meshwork, orbital and sclera stem cells have properties identical to cells of mesenchymal origin but their potential has yet to be experimentally determined and validated. Retinal and retinal pigment epithelium stem cells remain the most sought out stem cells for curing retinal degenerative disorders, although treatments using them have resulted in variable outcomes. The functional aspects of the therapeutic application of lenticular stem cells are not known and need further attention. Recently, embryonic stem cell-derived retinal pigment epithelium has been used for treating patients with Stargardts disease and age-related macular degeneration. Overall, the different stem cells residing in different components of the eye have shown some success in clinical and animal studies in the field of regenerative medicine.

Stem cells hold promise for treating a wide variety of diseases, including degenerative disorders of the eye. The eye is an ideal organ for stem cell therapy because of its relative immunological privilege, surgical accessibility, and its being a self-contained system. The eye also has many potential target diseases amenable to stem cell-based treatment, such as corneal limbal stem cell deficiency, glaucoma, age-related macular degeneration (AMD), and retinitis pigmentosa (RP). Among them, AMD and glaucoma are the two most common diseases, affecting over 200 million people worldwide. Recent results on the clinical trial of retinal pigment epithelial (RPE) cells from human embryonic stem cells (hESCs) and induced pluripotent stem cells (iPSCs) in treating dry AMD and Stargardt’s disease in the US, Japan, England, and China have generated great excitement and hope. This marks the beginning of the ocular stem cell therapy era. The recent Zhongshan Ophthalmic Center Ocular Stem Cell Symposium discussed the potential applications of various stem cell types in stem cell-based therapies, drug discoveries and tissue engineering for treating ocular diseases.

EMBO Reports (2019) e48172 Stem cell‐based regenerative medicine has experienced turbulent times in recent years that have involved several scandals implicating prominent institutions and numerous retractions of papers from leading journals. But it has also seen real progress being made on the way to the clinic. It reflects the tension between human ambition and hope on one side and the need for hard evidence and solid data on the other side in a field where early expectations ran far ahead of reality. This tension may help explain the recent scandal involving stem cell surgeon Paolo Macchiarini, who was found guilty of scientific misconduct regarding an article in The Lancet , which has since been retracted. It is an ongoing saga as Macchiarini is still publishing in leading journals, even though his reputation as a surgeon and stem cell researcher has been compromised. > … the whole field of regenerative medicine had been tainted by a number of cases where […] researchers had taken short cuts and neglected basic science. The story dates back to 2004, when Macchiarini's team extracted some pig intestine and removed the cells to leave a collagen matrix that was then repopulated with muscle and fibroblasts to generate connective epidermal tissue. The patch was implanted in the airway of a 58‐year‐old patient suffering from respiratory problems after surgery for lung cancer [1]. Macchiarini then expanded this technique to whole tracheas using plastic scaffolds in addition to natural tissue from human donors. In another clinical case, the bare plastic scaffold was populated with stem cells taken from the patient's own bone marrow and implanted in a young female patient in 2008. This propelled Macchiarini almost to rock star status and an appointment by the Karolinska Institute, where he refined his technique by replacing donor windpipes with plastic …

Stem cells can be defined as cells that have the capacity to self-renew and the ability to generate differentiated progeny or multiple cell lineages. True stem cells can turn into any type of cells, while progenitor cells are more or less committed to becoming cell types of a particular tissue. Human corneal epithelial stem cells (CESCs) represent a great example and model of adult stem or progenitor cells. Human CESCs have been identified to locate in the basal epithelial layer of the limbus, and thus also referred as to limbal stem cells. We would like to use the both terms, stem and progenitor cells in this chapter based on previous use in the literature for more than two decades. Although the CESCs have been identified to reside at the limbus and many stem cell markers have been proposed, there is no consensus to date regarding the definitive markers for CESCs, and identification and isolation of these cells are still challenging. Based on evaluation of a variety of proposed markers, we have characterized that the CESCs located in the basal layer of human limbal epithelium are small primitive cells expressing three patterns of molecular markers, which represent a unique phenotype of putative corneal epithelial stem or progenitor cells. Based on adult stem cell criteria and the putative limbal stem cell phenotype, our group has attempted to enrich for human CESCs through novel approaches including cell-sizing, adhering to extracellular matrix collagen type IV, and cell sorting for side population or for expression of ABCG2 or connexin 43 cell surface markers. The 5 clonogenic populations isolated from limbal epithelium and its cultures by different methods show the properties that are characteristics of adult stem/progenitor cells: 1) relatively undifferentiated, 2) high proliferative potential, 3) self-renewal. Expansion and cultivation of corneal epithelial progenitor cells have been achieved using different methods, such as limbal tissue explant culture, and limbal epithelial cell suspension co-culture with mouse 3T3 fibroblast feed layer. To avoid the use of xeno-components, two cell lines of commercial human fibroblasts have been identified that support human corneal epithelial regeneration, and have potential use in replacing mouse 3T3 cells for corneal tissue bioengineering. The concept of CESCs has formed the basis for identifying a class of blinding diseases that display features of corneal epithelial stem cell deficiency or limbal stem cell deficiency (LSCD), where the limbal epithelium is damaged. LSCD is characterized by persistent or recurrent epithelial defects, ulceration, corneal vascularization, chronic inflammation, scarring, and conjunctivalization (conjunctival epithelial ingrowth). Only transplantation of CESCs can restore vision. Due to an increasing shortage of corneal donors, corneal tissue engineering is becoming an important discipline that holds great promise for corneal reconstruction. CESCs and optical substrates are known to be the most important factors for corneal tissue bioengineering in regenerative medicine. Our team has recently explored the utilization of natural donor corneal stroma in corneal tissue engineering. In combination with fresh limbal epithelium containing stem cells, and the donor corneal stroma, a great source of natural optical substrate, we developed a native-like corneal equivalent construct with proliferative potential. This corneal construct provides a new clinical cell therapy for corneal reconstruction.

To highlight the progress and future direction of limbal stem cell (LSC) therapies for the treatment of limbal stem cell deficiency (LSCD). Direct LSC transplantation have demonstrated good long-term outcomes. Cultivated limbal epithelial transplantation (CLET) has been an alternative to treat severe to total LSCD aiming to improve the safety and efficacy of the LSC transplant. A prospective early-stage uncontrolled clinical trial shows the feasibility and safety of CLET manufactured under xenobiotic free conditions. Other cell sources for repopulating of the corneal epithelium such as mesenchymal stem cells (MSCs) and induced pluripotent stem cells are being investigated. The first clinical trials of using MSCs showed short-term results, but long-term efficacy seems to be disappointing. A better understanding of the niche function and regulation of LSC survival and proliferation will lead to the development of medical therapies to rejuvenate the residual LSCs found in a majority of eyes with LSCD in vivo. Prior efforts have been largely focused on improving LSC transplantation. Additional effort should be placed on improving the accuracy of diagnosis and staging of LSCD, and implementing standardized outcome measures which enable comparison of efficacy of different LSCD treatments for different severity of LSCD. The choice of LSCD treatment will be customized based on the severity of LSCD in the future. New approaches for managing different stages of LSCD are being developed. This concise review summarizes the progresses in LSC therapies for LSCD, underlying mechanisms, limitations, and future areas of development.

Embryonic stem-cell-derived retinal pigment epithelial cells for macular degeneration

Background Stem cell transplantation is a promising therapy for degenerative retinal diseases, including age‐related macular degeneration (AMD), retinitis pigmentosa (RP), and Stargardt's disease (STGD). This study quantitatively evaluates long‐term outcomes of different stem cell treatments. Methods A systematic review and outcome prediction analysis were conducted using data published before September 1, 2024. The primary outcome was the change in best‐corrected visual acuity (BCVA), expressed as logarithm of the minimum angle of resolution (LogMAR), adjusted for baseline BCVA. Predictions were disease‐specific for dry AMD, wet AMD, RP, and STGD, refined through regression analysis while ensuring data standardization across studies. Results A meta‐analysis of 43 studies (666 eyes) revealed varying BCVA improvements at 6 months post‐treatment. For dry AMD, adipose‐derived mesenchymal stem cells (ADMSCs) showed the best improvement (0.65, 95% CI: 0.57–0.72 logMAR), exceeding that of human embryonic stem cells (hESCs, 0.49, 95% CI: 0.33–0.65 logMAR). For wet AMD, hESCs improved BCVA by 0.45 logMAR (95% CI: 0.28–0.61 logMAR). For RP, Wharton's jelly‐derived mesenchymal stem cells (WJMSCs) achieved the highest improvement (0.50, 95% CI: 0.28–0.73 logMAR), while umbilical cord‐derived mesenchymal stem cells (UCMSCs, 0.39, 95% CI: 0.31–0.46 logMAR) showed moderate effects. For STGD, ADMSCs were most effective (0.53, 95% CI: 0.42–0.64 logMAR). Conclusions Stem cell therapy demonstrates significant potential for treating AMD, RP, and STGD, with ADMSCs showing superior benefits in dry AMD and STGD, hESCs being more effective in wet AMD, and WJMSCs outperforming other therapies in RP. These comparative insights highlight disease‐specific advantages of different stem cell sources, informing future clinical applications.

Regenerative treatment of ophthalmic diseases with stem cells: Principles, progress, and challenges

There is a rise in the number of people who have vision loss due to retinal diseases, and conventional therapies for treating retinal degeneration fail to repair and regenerate the damaged retina. Several studies in animal models and human trials have explored the use of stem cells to repair the retinal tissue to improve visual acuity. In addition to the treatment of age-related macular degeneration (AMD) and diabetic retinopathy (DR), stem cell therapies were used to treat genetic diseases such as retinitis pigmentosa (RP) and Stargardt’s disease, characterized by gradual loss of photoreceptor cells in the retina. Transplantation of retinal pigment epithelial (RPE) cells derived from embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) have shown promising results in improving retinal function in various preclinical models of retinal degeneration and clinical studies without any severe side effects. Mesenchymal stem cells (MSCs) were utilized to treat optic neuropathy, RP, DR, and glaucoma with positive clinical outcomes. This review summarizes the preclinical and clinical evidence of stem cell therapy and current limitations in utilizing stem cells for retinal degeneration.

A primary pathological defect in the heritable eye disorder Stargardt disease is excessive accumulation of cytotoxic lipofuscin bisretinoids in the retina. Age-dependent accumulation of lipofuscin in the retinal pigment epithelium (RPE) matches the age-dependent increase in the incidence of the atrophic (dry) form of age-related macular degeneration (AMD) and therefore may be one of several pathogenic factors contributing to AMD progression. Lipofuscin bisretinoid synthesis in the retina depends on the influx of serum retinol from the circulation into the RPE. Formation of the tertiary retinol-binding protein 4 (RBP4)-transthyretin-retinol complex in the serum is required for this influx. Herein, we report the pharmacological effects of the non-retinoid RBP4 antagonist, BPN-14136. BPN-14136 dosing in the Abca4-/- mouse model of increased lipofuscinogenesis significantly reduced serum RBP4 levels and inhibited bisretinoid synthesis, and this inhibition correlated with a partial reduction in visual cycle retinoids such as retinaldehydes serving as bisretinoid precursors. BPN-14136 administration at doses inducing maximal serum RBP4 reduction did not produce changes in the rate of the visual cycle, consistent with minimal changes in dark adaptation. Abca4-/- mice exhibited dysregulation of the complement system in the retina, and BPN-14136 administration normalized the retinal levels of proinflammatory complement cascade components such as complement factors D and H, C-reactive protein, and C3. We conclude that BPN-14136 has several beneficial characteristics, combining inhibition of bisretinoid synthesis and reduction in retinaldehydes with normalization of the retinal complement system. BPN-14136, or a similar compound, may be a promising drug candidate to manage Stargardt disease and dry AMD.

At this time, the main targets of stem cell therapy for retinal degenerative disease are age-related macular degeneration, Stargardt disease, and retinitis pigmentosa. The goal of stem cell therapy is to either to “rescue” the surviving retinal cells (by providing the necessary support or generating neurotrophic agents) and/or to “replace” the cells that have degenerated. Stem cells being used in ongoing early human trials to treat degenerative retinal disease include induced pluripotent stem cell (iPSC)-derived retinal pigment epithelium (RPE), embryonic stem cell-derived RPE, iPSC-neural progenitor cells, bone marrow-derived stem cells, and human central nervous system derived stem cells among others. It is too early to judge the outcome of these sources of tissue, but early results are positive. Continuing research in various aspects of transplantation- establishing cell lines without danger of tumor formation or immune rejection, refining surgical techniques and instruments, and identifying factors that promote cell survival, differentiation, and integration of the transplanted cells, should allow for rapid and continued progress in the field.

Limbal stem-cell deficiency (LSCD) is a disease characterized by persistent or recurrent epithelial defects, chronic inflammation, and conjunctiva migration onto the cornea (conjunctivalization) with severe visual impairment. Lamellar and/or penetrating keratoplasty cannot be successful as donor corneal epithelium is replaced by that of the recipient within months. In the presence of LSCD graft re-epithelialisation will not take place, with subsequent recurrence of conjunctivalization and graft failure. Stem-cell transplantation to treat LSCD is a step in the reconstruction of the ocular surface, while lamellar or penetrating corneal grafting will finally restore corneal transparency, leading to the recovery of visual capacity. The source of stem cells is typically classified as autologous and allogeneic. Unilateral or partial bilateral LSCDs can be treated with autologous limbal stem cell transplantation, while total bilateral deficiency requires allogeneic LSCs, or other sources of autologous cells such as oral epithelial stem cells.

Translating Stem Cell Studies to the Clinic for CNS Repair: Current State of the Art and the Need for a Rosetta Stone