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Purpose: The aim of this study was to evaluate how Scheimpflug-derived parameters of eyes with Fuchs endothelial corneal dystrophy (FECD) are influenced by Descemet membrane endothelial keratoplasty (DMEK) depending on FECD severity and the presence of subclinical edema., Methods: A retrospective cohort study including 115 eyes (115 patients) that underwent DMEK for FECD and a control group of 27 eyes with nonpathological corneas was conducted. Preoperative and 6 months postoperative Scheimpflug imaging was used to analyze pachymetry, presence of tomographic features (loss of isopachs/displacement of the thinnest point/focal posterior depression), and corneal backscatter. FECD severity was based on the modified Krachmer scale and the absence/presence of subclinical edema., Results: Scheimpflug-derived pachymetry, tomographic, and corneal backscatter parameters were correlated with FECD severity, and all changed from preoperatively to postoperatively (all P < 0.05). Postoperative central corneal thickness, anterior and posterior corneal backscatter, and presence of focal posterior depression remained different from the control group (all P < 0.05). Of eyes without preoperative clinical edema (n = 75), 18.7% showed 0 or 1 tomographic feature (no edema group) and 82.4% had 2 or 3 features (subclinical edema group). Compared with the control group, postoperative best-corrected visual acuity for the "no edema" group did not differ (0.03 ± 0.12 vs. -0.02 ± 0.08 logarithm of the minimum angle of resolution, P = 0.150) but was worse for the subclinical edema group (0.06 ± 0.08 vs. -0.02 ± 0.08 logarithm of the minimum angle of resolution, P = 0.001)., Conclusions: For eyes without preoperative edema, more parameters reversed back to 'normal' levels than for eyes with (sub)clinical edema. Although most analyzed parameters correlated with FECD severity, corneal tomography might be best suited for objective grading of disease severity to aid in surgical decision-making., Competing Interests: G. R. J. Melles is a consultant for DORC International/Dutch Ophthalmic USA and SurgiCube International. The remaining authors have no funding or conflicts of interest to disclose., (Copyright © 2022 Wolters Kluwer Health, Inc. All rights reserved.)

In this study we describe peripheral corneal endothelial cell migration in vitro in the absence and presence of a ROCK-inhibitor. For this study, 21 corneal endothelial graft rims, with attached trabecular meshwork (TM), were prepared from Descemet membrane-endothelial cell sheets by 6.5 mm trepanation. For the initial proof-of-concept, 7 outer graft rims were cultured in a thermo-reversible hydrogel matrix for up to 47 days. To assess the effect of a ROCK-inhibitor, 14 paired outer rims were cultured either with or without ROCK-inhibitor for up to 46 days. At the end of culture, tissue was retrieved from the hydrogel matrix and examined for cell viability and expression of different endothelial cell markers (ZO-1, Na+/K+-ATPase, NCAM, glypican, and vimentin). All cultured rims remained viable and displayed either single regions (n = 5/21) or collective areas (n = 16/21) of cell migration, regardless of the presence or absence of ROCK-inhibition. Migration started after 4±2 days and continued for at least 29 days. The presence of ROCK-inhibitor seemed to contribute to a more regular cell morphology of migrating cells. In addition, 7 outer rims demonstrated a phenotypically distinct late-onset but fast-growing cell population emerging from the area close to the limbus. These cells emerged after 3 weeks of culture and appeared less differentiated compared to other areas of migration. Immunostaining showed that migrated cells maintained the expression patterns of endothelial cell markers. In conclusion, we observed 2 morphologically distinct migrating cell populations with the first type being triggered by a broken physical barrier, which disrupted contact inhibition and the second, late-onset type showing a higher proliferative capacity though appearing less differentiated. This cell subpopulation appeared to be mediated by stimuli other than loss of contact inhibition and ROCK-inhibitor presence. Further exploration of the differences between these cell types may assist in optimizing regenerative treatment options for endothelial diseases., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Miron et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Over the past twenty years, there have been many developments in the treatment of corneal diseases, especially in the field of corneal transplantation. By targeting treatments solely to the affected layers of the cornea, the procedures have become less invasive and the cornea recovers significantly faster. Despite the success of these new surgical techniques, new (cell) regenerative therapies are emerging.

The corneal endothelium located on the posterior corneal surface is responsible for regulating stromal hydration. This is contributed by a monolayer of corneal endothelial cells (CECs), which are metabolically active in a continuous fluid-coupled efflux of ions from the corneal stroma into the aqueous humor, preventing stromal over-hydration and preserving the orderly arrangement of stromal collagen fibrils, which is essential for corneal transparency. Mature CECs do not have regenerative capacity and cell loss due to aging and diseases results in irreversible stromal edema and a loss of corneal clarity. The current gold standard of treatment for this worldwide blindness caused by corneal endothelial failure is the corneal transplantation using cadaveric donor corneas. The top indication is Fuchs corneal endothelial dystrophy/degeneration, which represents 39% of all corneal transplants performed. However, the global shortage of transplantable donor corneas has restricted the treatment outcomes, hence instigating a need to research for alternative therapies. One such avenue is the CEC regeneration from endothelial progenitors, which have been identified in the peripheral endothelium and the adjacent transition zone. This review examines the evidence supporting the existence of endothelial progenitors in the posterior limbus and summarizes the existing knowledge on the microanatomy of the transitional zone. We give an overview of the isolation and ex vivo propagation of human endothelial progenitors in the transition zone, and their growth and differentiation capacity to the corneal endothelium. Transplanting these bioengineered constructs into in vivo models of corneal endothelial degeneration will prove the efficacy and viability, and the long-term maintenance of functional endothelium. This will develop a novel regenerative therapy for the management of corneal endothelial diseases.

Fuchs endothelial corneal dystrophy (FECD) is amongst one of the most common indications for endothelial keratoplasty worldwide. Despite being originally described among Caucasians, it is now known to be prevalent among a large number of populations, including Asians. While the FECD phenotype is classically described as that of central guttate and pigment deposits associated with corneal endothelial dysfunction, there are subtle yet important differences in how FECD and its phenocopies may present in Caucasians vs Asians. Such differences are paralled by genotypic variations and disease management preferences which appear to be geographically and ethnically delineated. This article provides a succinct review of such differences, with a focus on diagnostic and management issues which may be encountered by ophthalmologists practicing in the different geographic regions, when evaluating a patient with FECD.

Purpose of Review: To provide an update on the state of development of novel therapeutic modalities for the treatment of corneal diseases., Recent Findings: Novel corneal therapeutics may be broadly classified as cell therapy, regenerative medicine, bioengineered corneal grafts and gene therapy. Cell therapy encompasses cultivation of cells, such as corneal endothelial cells (CECs) and keratocytes to replenish the depleted native cell population. Regenerative medicine is mainly applicable to the corneal endothelium, and is dependent on the ability of native, healthy CECs to restore the corneal endothelium following trauma or descemetorhexis; this approach may be effective for the treatment of Peter's anomaly and Fuchs endothelial corneal dystrophy (FECD). Bioengineered corneal grafts are synthetic constructs designed to replace cadaveric corneal grafts; tissue-engineered endothelial-keratoplasty grafts and bioengineered stromal grafts have been experimented in animal models with favourable results. Gene therapy with antisense oligonucleotide and CRISPR endonucleases, including deactivated Cas9, may potentially be used to treat FECD and TGFBI-related corneal dystrophies., Summary: These novel therapeutic modalities may potentially supersede keratoplasty as the standard of care in the future.

(1) Rho-associated coiled-coil protein kinase (ROCK) signaling cascade impacts a wide array of cellular events. For cellular therapeutics, scalable expansion of primary human corneal endothelial cells (CECs) is crucial, and the inhibition of ROCK signaling using a well characterized ROCK inhibitor (ROCKi) Y-27632 had been shown to enhance overall endothelial cell yield. (2) In this study, we compared several classes of ROCK inhibitors to both ROCK-I and ROCK-II, using in silico binding simulation. We then evaluated nine ROCK inhibitors for their effects on primary CECs, before narrowing it down to the two most efficacious compounds—AR-13324 (Netarsudil) and its active metabolite, AR-13503—and assessed their impact on cellular proliferation in vitro. Finally, we evaluated the use of AR-13324 on the regenerative capacity of donor cornea with an ex vivo corneal wound closure model. Donor-matched control groups supplemented with Y-27632 were used for comparative analyses. (3) Our in silico simulation revealed that most of the compounds had stronger binding strength than Y-27632. Most of the nine ROCK inhibitors assessed worked within the concentrations of between 100 nM to 30 µM, with comparable adherence to that of Y-27632. Of note, both AR-13324 and AR-13503 showed better cellular adherence when compared to Y-27632. Similarly, the proliferation rates of CECs exposed to AR-13324 were comparable to those of Y-27632. Interestingly, CECs expanded in a medium supplemented with AR-13503 were significantly more proliferative in (i) untreated vs. AR-13503 (1 μM; * p < 0.05); (ii) untreated vs. AR-13503 (10 μM; *** p < 0.001); (iii) Y-27632 vs. AR-13503 (10 μM; ** p < 0.005); (iv) AR-13324 (1 μM) vs. AR-13503 (10 μM; ** p < 0.005); and (v) AR-13324 (0.1 μM) vs. AR-13503 (10 μM; * p < 0.05). Lastly, an ex vivo corneal wound healing study showed a comparable wound healing rate for the final healed area in corneas exposed to Y-27632 or AR-13324. (4) In conclusion, we were able to demonstrate that various classes of ROCKi compounds other than Y-27632 were able to exert positive effects on primary CECs, and systematic donor-match controlled comparisons revealed that the FDA-approved ROCK inhibitor, AR-13324, is a potential candidate for cellular therapeutics or as an adjunct drug in regenerative treatment for corneal endothelial diseases in humans. [ABSTRACT FROM AUTHOR]

Corneal transplantation is currently the most effective treatment to restore corneal clarity in patients with endothelial disorders. Endothelial transplantation, either by Descemet membrane endothelial keratoplasty (DMEK) or by Descemet stripping (automated) endothelial keratoplasty (DS(A)EK), is a surgical approach that replaces diseased Descemet membrane and endothelium with tissue from a healthy donor eye. Its application, however, is limited by the availability of healthy donor tissue. To increase the pool of endothelial grafts, research has focused on developing new treatment options as alternatives to conventional corneal transplantation. These treatment options can be considered as either 'surgery‐based', that is tissue‐efficient modifications of the current techniques (e.g. Descemet stripping only (DSO)/Descemetorhexis without endothelial keratoplasty (DWEK) and Quarter‐DMEK), or 'cell‐based' approaches, which rely on in vitro expansion of human corneal endothelial cells (hCEC) (i.e. cultured corneal endothelial cell sheet transplantation and cell injection). In this review, we will focus on the most recent developments in the field of the 'cell‐based' approaches. Starting with the description of aspects involved in the isolation of hCEC from donor tissue, we then describe the different natural and bioengineered carriers currently used in endothelial cell sheet transplantation, and finally, we discuss the current 'state of the art' in novel therapeutic approaches such as endothelial cell injection. [ABSTRACT FROM AUTHOR]

Since its development more than 15 years ago (Melles et al. 2006), Descemet membrane endothelial keratoplasty (DMEK) has become the preferred treatment option for endothelial diseases (Birbal et al. 2020). Misconception: A pseudophakic eye is easier to operate than a phakic eye Pseudophakic eyes enable DMEK-graft manipulation within a deeper AC than phakic eyes, without the risk of crystalline lens touch. Misconception: A loose roll is easier than a tight roll The preferable method for DMEK surgery is insertion of the DMEK-graft in a double-roll configuration into the anterior chamber (AC; Dapena et al. 2011). [Extracted from the article]

Fuchs endothelial corneal dystrophy (FECD) is the most common primary corneal endothelial dystrophy and the leading indication for corneal transplantation worldwide. FECD is characterized by the progressive decline of corneal endothelial cells (CECs) and the formation of extracellular matrix (ECM) excrescences in Descemet's membrane (DM), called guttae, that lead to corneal edema and loss of vision. FECD typically manifests in the fifth decades of life and has a greater incidence in women. FECD is a complex and heterogeneous genetic disease where interaction between genetic and environmental factors results in cellular apoptosis and aberrant ECM deposition. In this review, we will discuss a complex interplay of genetic, epigenetic, and exogenous factors in inciting oxidative stress, auto(mito)phagy, unfolded protein response, and mitochondrial dysfunction during CEC degeneration. Specifically, we explore the factors that influence cellular fate to undergo apoptosis, senescence, and endothelial-to-mesenchymal transition. These findings will highlight the importance of abnormal CEC-DM interactions in triggering the vicious cycle of FECD pathogenesis. We will also review clinical characteristics, diagnostic tools, and current medical and surgical management options for FECD patients. These new paradigms in FECD pathogenesis present an opportunity to develop novel therapeutics for the treatment of FECD. [ABSTRACT FROM AUTHOR]

Donor corneas with low endothelial cell densities (ECD) are deemed unsuitable for corneal endothelial transplantation. This study evaluated a two-step incubation and dissociation harvesting approach to isolate single corneal endothelial cells (CECs) from donor corneas for corneal endothelial cell-injection (CE-CI) therapy. To isolate CECs directly from donor corneas, optimization studies were performed where donor Descemet's membrane/corneal endothelium (DM/CE) were peeled and incubated in either M4-F99 or M5-Endo media before enzymatic digestion. Morphometric analyses were performed on the isolated single cells. The functional capacities of these cells, isolated using the optimized simple non-cultured endothelial cells (SNEC) harvesting technique, for CE-CI therapy were investigated using a rabbit bullous keratopathy model. The two control groups were the positive controls, where rabbits received cultured CECs, and the negative controls, where rabbits received no CECs. Whilst it took longer for CECs to dislodge as single cells following donor DM/CE incubation in M5-Endo medium, CECs harvested were morphologically more homogenous and smaller compared to CECs obtained from DM/CE incubated in M4-F99 medium (p < 0.05). M5-Endo medium was hence selected as the DM/CE incubation medium prior to enzymatic digestion to harvest CECs for the in vivo cell-injection studies. Following SNEC injection, mean central corneal thickness (CCT) of rabbits increased to 802.9 ± 147.8 μm on day 1, gradually thinned, and remained clear with a CCT of 385.5 ± 38.6 μm at week 3. Recovery of corneas was comparable to rabbits receiving cultured CE-CI (p = 0.40, p = 0.17, and p = 0.08 at weeks 1, 2, and 3, respectively). Corneas that did not receive any cells remained significantly thicker compared to both SNEC injection and cultured CE-CI groups (p < 0.05). This study concluded that direct harvesting of single CECs from donor corneas for SNEC injection allows the utilization of donor corneas unsuitable for conventional endothelial transplantation. [ABSTRACT FROM AUTHOR]