Corneal Endothelial Diseases Research Articles

DMEK graft: One size does not fit all.

Descemet membrane endothelial keratoplasty (DMEK) is a popular procedure for the treatment of corneal endothelial diseases mainly targeting Fuchs endothelial corneal dystrophy (FECD) and pseudophakic bullous keratopathy (PBK). Although DMEK has multiple advantages, it is challenging in terms of graft preparation and delivery. One of the crucial factors of DMEK graft preparation is determining the size of the graft. Evaluating risks and benefits of transplanting larger or smaller grafts compared with the descemetorhexis performed following a standard DMEK procedure thus becomes important. Advanced techniques like pre-loaded DMEK requires pre-selection of graft diameter without physical examination of the eye making it more challenging. Therefore, recognizing the benefits of graft size and the number of transplanted endothelial cells becomes essential. Smaller DMEK grafts have been preferred and accepted for grafting. Larger diameter grafts have advantages but can be challenging due to higher detachment rates. We thus aim to review the challenges of preparing and delivering DMEK tissues with small or large diameter based on selected descemetorhexis area, discuss the outcomes based on different graft sizes, highlight related complications and suggest which cases may benefit from adopting smaller or larger graft size.

Cell therapy in corneal endothelial disease.

Endothelial keratoplasty is the current gold standard for treating corneal endothelial diseases, achieving excellent visual outcomes and rapid rehabilitation. There are, however, severe limitations to donor tissue supply and uneven access to surgical teams and facilities across the globe. Cell therapy is an exciting approach that has shown promising early results. Herein, we review the latest developments in cell therapy for corneal endothelial disease. We highlight the work of several groups that have reported successful functional outcomes of cell therapy in animal models, with the utilization of human embryonic stem cells, human-induced pluripotent stem cells and cadaveric human corneal endothelial cells (CECs) to generate populations of CECs for intracameral injection. The use of corneal endothelial progenitors, viability of cryopreserved cells and efficacy of simple noncultured cells, in treating corneal decompensation is of particular interest. Further additions to the collective understanding of CEC physiology, and the process of cultivating and administering effective cell therapy are reviewed as well. The latest developments in cell therapy for corneal endothelial disease are presented. The continuous growth in this field gives rise to the hope that a viable solution to the large numbers of corneal blind around the world will one day be reality.

Long-term observation after transplantation of cultured human corneal endothelial cells for corneal endothelial dysfunction

BackgroundCorneal transplantation is the only way to treat serious corneal diseases caused by corneal endothelial dysfunction. However, the shortage of donor corneal tissues and human corneal endothelial cells (HCECs) remains a worldwide challenge. We cultivated HCECs by the use of a conditioned medium from orbital adipose-derived stem cells (OASC-CM) in vitro. Then the HCECs were used to treat animal corneal endothelial dysfunction models via cell transplantation. The purpose of this study was to conduct a long-term observation and evaluation after cell transplantation.MethodsOrbital adipose-derived stem cells (OASCs) were isolated to prepare the conditioned medium (CM). HCECs were cultivated and expanded by the usage of the CM (CM-HCECs). Then, related corneal endothelial cell (CEC) markers were analyzed by immunofluorescence. The cell proliferation ability was also tested. CM-HCECs were then transplanted into monkey corneal endothelial dysfunction models by injection. We carried out a 24-month postoperative preclinical observation and verified the long-term effect by histological examination and transcriptome sequencing.ResultsCM-HCECs strongly expressed CEC-related markers and maintained polygonal cell morphology even after 10 passages. At 24 months after cell transplantation, there was a CEC density of more than 2400 cells per square millimeter (range, 2408–2685) in the experimental group. A corneal thickness (CT) of less than 550 μm (range, 490–510) was attained. Gene sequencing showed that the gene expression pattern of CM-HCECs was similar to that of transplanted cells and HCECs.ConclusionsTransplantation of CM-HCECs into monkey corneal endothelial dysfunction models resulted in a transparent cornea after 24 months. This research provided a promising prospect of cell-based therapy for corneal endothelial diseases.

Alternatives to endokeratoplasty: an attempt towards reducing global demand ofhuman donor corneas.

The cornea is an anterior transparent tissue of the eye that enables the transmission of surrounding light to the back of the eye, which is essential for maintaining clear vision. Corneal endothelial diseases can lead to partial or total blindness; hence, surgical replacement of the diseased corneal tissue with a healthy cadaveric donor graft becomes necessary when the endothelium is damaged. Keratoplasties face a huge challenge due to a worldwide shortage in the supply of human donor corneas. Hence, alternative solutions such as cell or tissue engineering-based therapies have been investigated for reducing the global demand ofdonor corneas. This review aims at highlighting studies that have been successful at replacing partial or total endothelial keratoplasty.

Approaches for corneal endothelium regenerative medicine

The state of the art therapy for treating corneal endothelial disease is transplantation. Advances in the reproducibility and accessibility of surgical techniques are increasing the number of corneal transplants, thereby causing a global deficit of donor corneas and leaving 12.7 million patients with addressable visual impairment. Approaches to regenerate the corneal endothelium offer a solution to the current tissue scarcity and a treatment to those in need. Methods for generating corneal endothelial cells into numbers that could address the current tissue shortage and the possible strategies used to deliver them have now become a therapeutic reality with clinical trials taking place in Japan, Singapore and Mexico. Nevertheless, there is still a long way before such therapies are approved by regulatory bodies and become clinical practice. Moreover, acellular corneal endothelial graft equivalents and certain drugs could provide a treatment option for specific disease conditions without the need of donor tissue or cells. Finally, with the emergence of gene modulation therapies to treat corneal endothelial disease, it would be possible to treat presymptomatic patients or those presenting early symptoms, drastically reducing the need for donor tissue. It is necessary to understand the most recent developments in this rapidly evolving field to know which conditions could be treated with which approach. This article provides an overview of the current and developing regenerative medicine therapies to treat corneal endothelial disease and provides the necessary guidance and understanding towards the treatment of corneal endothelial disease.

Effect of Physiological Oxygen on Primary Human Corneal Endothelial Cell Cultures.

PurposePrimary human corneal endothelial cells (HCEnCs) cultured in room air are exposed to significantly higher O2 concentrations [O2] than what is normally present in the eye. We evaluated the growth and metabolism of HCEnCs cultured under physiological [O2] (2.5%; [O2]2.5) and room air ([O2]A).MethodsPrimary cultures of HCEnCs from normal donors and donors with Fuchs dystrophy were grown at [O2]2.5 and [O2]A. Growth and morphology were compared using phase-contrast microscopy, zonula occludens (ZO-1) localization, cell density measurements, and senescence marker staining. CD44 (cell quality) and HIF-1α (hypoxia-inducible factor-1α) levels were evaluated by Western blotting. Cell adaptability to a reversal of [O2] growth conditions was measured with cell viability assays, and cell metabolism was assessed via oxygen consumption and extracellular acidification rates.ResultsHCEnCs grown at [O2]A and [O2]2.5 displayed similar morphologies, ZO-1 localization, CD44 expression, and senescence. Cells from donors with Fuchs dystrophy grew better at [O2]2.5 than at [O2]A. HIF-1α was undetectable. Cells displayed greater viability at [O2]2.5 than at [O2]A. HCEnCs showed significantly greater proton leak (P < 0.01), nonmitochondrial oxygen consumption (P < 0.01), and spare capacity (P < 0.05) for oxygen consumption rates, and greater basal glycolysis (P < 0.05) with a decreased glycolytic reserve capacity (P < 0.05) for extracellular acidification rates.ConclusionsPrimary HCEnCs show unique metabolic characteristics at physiologic [O2]. The effect of [O2] for optimization of HCEnC culture conditions should be considered.Translational RelevanceWith the advance of cell-based therapeutics for corneal endothelial diseases, [O2] should be considered an important variable in the optimization of HCEnC culture conditions.

Corneal Densitometry After Uneventful Descemet Membrane Endothelial Keratoplasty-5-Year Outcomes.

The purpose of this study is to analyze the course of corneal densitometry (CD), endothelial cell density (ECD), best-corrected visual acuity (BCVA), and central corneal thickness (CCT) 5 years after uneventful Descemet membrane endothelial keratoplasty (DMEK). Sixty uneventful cases (51 patients) with a minimum follow-up of 5 years were included. CD of various corneal layers (anterior, central, posterior, and total layer) and zones (0-2 mm, 2-6 mm, and 6-10 mm) were measured with Scheimpflug tomography. ECD, BCVA, and CCT were also evaluated. Total CD at 0 to 2 mm and 2 to 6 mm zones significantly decreased from 33 ± 10 and 27.8 ± 8 grayscale units (GSU) preoperatively to 21.8 ± 3.1 and 22.2 ± 4.2 GSU at 5 years, respectively ( P < 0.001). On the contrary, total CD at the 6 to 10 mm zone significantly increased from 30 ± 8.3 GSU preoperatively to 34.6 ± 7.8 GSU at 5 years ( P < 0.001). ECD significantly decreased from 2496 ± 267 cells/mm 2 preoperatively to 1063 ± 470 cells/mm 2 at 5 years ( P < 0.001). Similarly, CCT significantly decreased from 686 ± 109 μm preoperatively to 557 ± 37 μm at 5 years ( P < 0.001). Postoperative BCVA was significantly better after DMEK for every examination time point. Despite a slight CD increase at all layers of all corneal zones from the second to the fifth postoperative year, the excellent visual outcome was maintained throughout 5-year follow-up. Thus, DMEK seems to effectively treat corneal endothelial disease in the long term.

Posterior lamellar keratoplasty: techniques, outcomes, and recent advances.

Over the past 2 decades, posterior lamellar keratoplasty (PLK) has emerged as an alternative to penetrating keratoplasty in the treatment of corneal endothelial disorders. The reasons for this trend include the search for a safer procedure to replace diseased endothelium that provides faster and better visual rehabilitation and reduces the need for postoperative care. Different surgical techniques, surgical instruments, devices, and lasers have been introduced to overcome technical difficulties, thus improving clinical outcomes. Yet, surgeons and eye banks must address the complications and limitations that arise during the transition to these new techniques. This review discusses the most significant aspects of the evolution of PLK, including a detailed description of current techniques and the direction of future treatment for corneal endothelial disease with the use of laser-assisted surgery, bioengineered corneas, cell therapy, and new pharmacologic therapy.

Aqueous humour cytokine profiles after Descemet\u2019s membrane endothelial keratoplasty

The aim of this study was to compare aqueous humour (AqH) cytokine profiles before and after Descemet’s membrane endothelial keratoplasty (DMEK) in eyes with bullous keratopathy (BK) and age-matched normal eyes. A total of 49 participants (mean age 75.0 ± 13.5 years) were divided into three groups: (1) BK patients scheduled for DMEK (BK group); (2) patients after DMEK (DMEK group; average postoperative time 1,018 ± 460 days); and (3) control participants without corneal endothelial disease scheduled for cataract surgery (control group). AqH (0.2 mL) was collected from all participants, and the levels of various AqH cytokines were simultaneously measured using a multiplex bead immunoassay. The DMEK group exhibited significantly lower concentrations of several pro-inflammatory cytokines, such as IL-1β, IL-5, IL-6, IL-10, and IL-8, and granulocyte colony stimulating factor than the BK group. In addition, the levels of IL-1β and IL-5 were significantly lower in the DMEK group than in the control group. The AqH levels of several pro-inflammatory cytokines were significantly reduced after DMEK, indicating that regeneration of the endothelial pump owing to the transplantation of a healthy endothelium might have a positive effect on anterior chamber inflammation.

Animal models of corneal endothelial dysfunction to facilitate development of novel therapies.

Progressive corneal endothelial disease eventually leads to corneal edema and vision loss due to the limited regenerative capacity of the corneal endothelium in vivo and is a major indication for corneal transplantation. Despite the relatively high success rate of corneal transplantation, there remains a pressing global clinical need to identify improved therapeutic strategies to address this debilitating condition. To evaluate the safety and efficacy of novel therapeutics, there is a growing demand for pre-clinical animal models of corneal endothelial dysfunction. In this review, experimentally induced, spontaneously occurring and genetically modified animal models of corneal endothelial dysfunction are described to assist researchers in making informed decisions regarding the selection of the most appropriate animal models to meet their research goals.

Activation of kelch-like ECH-associated protein 1/nuclear factor erythroid 2-related factor 2/antioxidant response element pathway by curcumin enhances the anti-oxidative capacity of corneal endothelial cells

Fuchs endothelial corneal dystrophy is one of the most common indications for corneal transplantation, and impaired anti-oxidative function is observed in corneal endothelial cells (CECs). Curcumin is well-known for its anti-oxidative property; but, no study has examined the effect of curcumin on anti-oxidative therapeutic roles in corneal endothelial disease. In our experiments, oxidative stress 0.25 mM tert-butyl hydroperoxide for 2 h was induced in immortalized human CECs pretreated with curcumin. Cell behavior and viability, reactive oxygen species production, and the protein expression of the kelch-like ECH-associated protein 1 (Keap1)/nuclear factor erythroid 2-related factor 2(Nrf2)/antioxidant response element (ARE) pathway were examined; the Keap1/Nrf2/ARE pathway is crucial anti-oxidative pathway of curcumin. The results showed that pretreatment with 12.5 μM curcumin significantly reduced the ROS production and improved the survival of CECs under oxidative stress. In addition, curcumin pretreatment significantly increased the expression of nuclear Nrf2, and the productions of superoxide dismutase 1 and heme oxygenase-1, which were the target anti-oxidative enzymes of the Keap1/Nrf2/ARE pathway. Our findings showed that curcumin enhanced the growth and differentiation of CECs under oxidative stress. The activation of Keap1/Nrf2/ARE pathway by curcumin was crucial for CECs to improve their anti-oxidative capacity.

New Therapies for Corneal Endothelial Diseases: 2020 and Beyond.

Penetrating keratoplasty used to be the only surgical technique for the treatment of end-stage corneal endothelial diseases. Improvements in surgical techniques over the past decade have now firmly established endothelial keratoplasty as a safe and effective modality for the treatment of corneal endothelial diseases. However, there is a worldwide shortage of corneal tissue, with more than 50% of the world having no access to cadaveric tissue. Cell injection therapy and tissue-engineered endothelial keratoplasty may potentially offer comparable results as endothelial keratoplasty while maximizing the use of cadaveric donor corneal tissue. Descemet stripping only, Descemet membrane transplantation, and selective endothelial removal are novel therapeutic modalities that take this a step further by relying on endogenous corneal endothelial cell regeneration, instead of allogenic corneal endothelial cell transfer. Gene therapy modalities, including antisense oligonucleotides and clustered regularly interspaced short palindromic repeats-based gene editing, offer the holy grail of potentially suppressing the phenotypic expression of genetically determined corneal endothelial diseases at the asymptomatic stage. We now stand at the crossroads of exciting developments in medical technologies that will likely revolutionize the way we treat corneal endothelial diseases over the next 2 decades.

Rho kinase (ROCK) inhibitors in the management of corneal endothelial disease.

Rho kinase (ROCK) inhibitors are growing increasingly relevant in ophthalmology, and the goal of this review is to summarize their mechanisms of action and potential applications in the subspecialties of glaucoma, retina, and cornea. We will focus specifically on corneal endothelial wound healing, for which ROCK inhibition demonstrates particular promise. ROCK inhibition has been shown to promote corneal endothelial cell proliferation, increase intercellular adhesion, and suppress apoptosis. Topical ROCK inhibitor treatment has exhibited potential use in Fuchs endothelial dystrophy, corneal edema from acute surgical trauma and other etiologies, and tissue engineering therapy for the endothelial disease. Ripasudil and netarsudil, the two ROCK inhibitors available for ophthalmic use, are generally very well tolerated with mild and transient local side effects. ROCK inhibitors are revolutionizing the subspecialty of cornea, and further research is needed to compare long-term outcomes of ROCK inhibitor therapy to those of conventional endothelial keratoplasty, including visual acuity and endothelial cell density. Other possible avenues include the use of ROCK inhibitors to prolong corneal graft survival, and early data appears promising.

Diseases of the corneal endothelium

The corneal endothelial monolayer and associated Descemet's membrane (DM) complex is a unique structure that plays an essential role in corneal function. Endothelial cells are neural crest derived cells that rest on a special extracellular matrix and play a major role in maintaining stromal hydration within a narrow physiologic range necessary for clear vision. A number of diseases affect the endothelial cells and DM complex and can impair corneal function and vision. This review addresses different human corneal endothelial diseases characterized by loss of endothelial function including: Fuchs endothelial corneal dystrophy (FECD), posterior polymorphous corneal dystrophy (PPCD), congenital hereditary endothelial dystrophy (CHED), bullous keratopathy, iridocorneal endothelial (ICE) syndrome, post-traumatic fibrous downgrowth, glaucoma and diabetes mellitus.

Translational and Regulatory Challenges of Corneal Endothelial Cell Therapy: A Global Perspective.

Cell therapies are emerging as a unique class of clinical therapeutics in medicine. In 2015, Holoclar (ex vivo expanded autologous human corneal epithelial cells containing stem cells) gained the regulatory approval for treating limbal stem cell deficiency after chemical eye burn. This has set a precedent in ophthalmology and in medicine, reinforcing the therapeutic promise of cell therapy. However, to generalize and commercialize cell therapies on a global scale, stringent translational and regulatory requirements need to be fulfilled at both local and international levels. Over the past decade, the Singapore group has taken significant steps in developing human corneal endothelial cell (HCEnC) therapy for treating corneal endothelial diseases, which are currently the leading indication for corneal transplantation in many countries. Successful development of HCEnC therapy may serve as a novel solution to the current global shortage of donor corneas. Based on the experience in Singapore, this review aims to provide a global perspective on the translational and regulatory challenges for bench-to-bedside translation of cell therapy. Specifically, we discussed about the characterization of the critical quality attributes (CQA), the challenges that can affect the CQA, and the variations in the regulatory framework embedded within different regions, including Singapore, Europe, and the United States. Impact statement Functional corneal endothelium is critical to normal vision. Corneal endothelial disease-secondary to trauma, surgery, or pathology-represents an important cause of visual impairment and blindness in both developed and developing countries. Currently, corneal transplantation serves as the current gold standard for treating visually significant corneal endothelial diseases, although limited by the shortage of donor corneas. Over the past decade, human corneal endothelial cell therapy has emerged as a promising treatment option for treating corneal endothelial diseases. To allow widespread application of this therapy, significant regulatory challenges will need to be systematically overcome.

Emerging Technologies to Solve the Key Issues in Endothelial Keratoplasty

Endothelial keratoplasty (EK) is increasingly used to treat corneal endothelial disease. However, issues remain with EK including a non-zero rejection rate, shortage of donor tissue, and technical challenges for Descemet’s membrane endothelial keratoplasty. Technologies that might solve these issues include small-molecule drugs, surgical innovations, cell therapy, tissue engineering, and gene therapy. Our purpose is to inform ophthalmologists, eye bankers, and eye health professionals of technologies that could impact practice in the near future. Recent advances are targeted at solving known issues with EK. Although none is yet routine, some are in clinical trials. Technologies that are closest to an ideal solution are those facing the highest regulatory and commercial hurdles. In the future, there will multiple options for the treatment of corneal endothelial cell failure. Which of these differing treatments will be used will depend on their efficacy as well as commercial, regulatory, and patient factors.

PAX6, modified by SUMOylation, plays a protective role in corneal endothelial injury

Treating corneal endothelial diseases tends to be challenging as human corneal endothelial cells (CECs) do not proliferate in vivo. The pathogenesis or mechanisms underlying injured CECs need further studies. The abnormal expression of PAX6, which is an essential transcription factor for corneal homeostasis, exhibits corneal endothelial defects. However, the effects of PAX6 protein involved in corneal endothelial wound process are still unknown. Here, we found the upregulated protein levels of PAX6 in human corneal endothelial monolayer after injury; the expression of PAX6 also increased in murine and rat corneal endothelium injury models. Enforced PAX6 expression could alleviate the damages to CECs via regulating permeability by prompting cellular tight junction. In addition, SUMOylation mainly happened on both K53 and K89 residues of 48-kD PAX6 (the longest and main isoform expressed in cornea), and de-SUMOylation promoted the stability of PAX6 protein in vitro. In CECs of SENP1+/− mice, increased SUMOylation levels leading to instability and low expression of PAX6, delayed the repair of CECs after injury. Furthermore, overexpression of PAX6 accelerated the rate of corneal endothelial repair of SENP1+/− mice. Our findings indicate that SENP1-mediated de-SUMOylation improving the stability of PAX6, amplifies the protective effects of PAX6 on corneal endothelial injuries, highlighting potentials of PAX6 and/or SUMOylation to be used as a treatment target for corneal endothelial disorders.

Bioengineering of Human Corneal Endothelial Cells from Single- to Four-Dimensional Cultures

To summarize the recent advances of clinical and preclinical studies for corneal endothelial tissue bioengineering. The challenges facing the generation of a clinical applicable corneal endothelial graft can be broadly classified into cell source selection, culture medium optimization, scaffold establishment, and the following three- and four-dimensional (4D) corneal construction. Based on the current advances in primary human corneal endothelial cell (HCEC) culture and good manufacturing practice (GMP)–compliant medium development, the first clinical trial of bioengineered HCEC injection therapy has been conducted with encouraging results. Other significant findings include the in vivo experiments of the stem cell–derived HCEC, the development of serum-, xeno-, and additive-free media, and the construction of 4D scaffold. It can be anticipated that tissue engineering–based therapy toward corneal endothelial diseases will replace the current keratoplasty method as a promising treatment option in the near future.

The Impact of Glaucoma Drainage Devices on the Cornea

To discuss the relevant clinical associations between glaucoma drainage device (GDD) implantation and health of the cornea, focusing on corneal endothelial disease and success of viability of keratoplasty. Corneal endothelial injury and risk of graft failure continues to demonstrate association with GDD placement; however, newer studies demonstrate improved outcomes associated with Descemet membrane endothelial keratoplasty. Additionally, newer adverse effects associated with GDD placement have been described. GDD implantation certainly carries risk of endothelial injury and keratoplasty viability, but newer keratoplasty techniques may improve visual outcomes. Furthermore, attention to placement and location of the GDD may affect the risk of progressive corneal injury.

Descemet membrane endothelial keratoplastyDMEK -Donor and recipient step by step

In 2020 Descemet membrane endothelial keratoplasty (DMEK) has become the gold standard in Germany for the treatment of corneal endothelial diseases; however, the widespread use of DMEK was initially limited due to problems with donor preparation and the difficulty of correctly and gently unfolding the endothelial Descemet membrane (EDM) in the anterior chamber. Following the situational donor selection, the safe single-handed donor preparation of the cornea without tearing or even rupturing the EDM, including the indispensable peripheral semicircular orientation marking. Also presented is a step by step atraumatic loading of the glass cartridge, the introduction of the EDM roll into the anterior chamber, and its safe step by step unfolding based on (1)the sequential use of jets of fluid, (2)repeated tapping on the peripheral/central cornea, (3)controlled flattening of the anterior chamber, (4)use of air bubbles of defined size and finally, the fixation of the EDM in correct orientation to the back of the host cornea with agas bubble. Since every wrong step with DMEK can have far-reaching consequences for patient and surgeon, this step by step pragmatic approach should minimize the incidence of donor tissue damage and failure in patient maneuvers.