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Corneal endothelium cells (CECs) regulate corneal hydration between the leaky barrier of the corneal endothelium and the ionic pumps on the surface of CECs. As CECs do not regenerate, loss of CECs leads to poor vision and corneal blindness. Corneal transplant is the only treatment option; however, there is a severe shortage of donor corneas globally. Cell therapy using propagated primary human CECs is an alternative approach to corneal transplantations, and proof of functionality is crucial for validating such CECs. Expression markers like Na-K-ATPase and ZO-1 are typical but not specific to CECs. Assessing the barrier function of the expanded CECs via electrical resistance (i.e., TEER and Ussing's chamber) involves difficult techniques and is thus impractical for clinical application. Calcium has been demonstrated to affect the paracellular permeability of the corneal endothelium. Its absence alters morphology and disrupts apical junctions in bovine CECs, underscoring its importance. Calcium signaling patterns such as calcium waves affect the rate of wound healing in bovine CECs. Therefore, observing calcium waves in expanded CECs could provide valuable insights into their health and functional integrity. Mechanical or chemical stimulations, combined with Ca2+-sensitive fluorescent dyes and time-lapse imaging, can be used to visualize these waves, which could potentially be used to qualify expanded CECs. [ABSTRACT FROM AUTHOR]

PurposeMethodsResultsConclusionTo analyze the changes of corneal endothelial cells in Chinese patients with unilateral Fuchs’ uveitis syndrome (FUS) and investigate the factors relevant to these changes.Bilateral specular microscopic examination was performed in 459 Chinese patients with unilateral FUS from April 2008 to April 2023. The affected eyes constituted the study group, while the contralateral eyes served as controls.The median values of endothelial cell density (ECD), cell count, total cell size, and hexagonality were significantly lower in the FUS eyes compared to the control eyes (p < 0.001). The median values of average cell size, maximum cell size, SD of cell size, and CV were significantly higher in the FUS eyes compared to the control eyes (p < 0.001). Central ECD showed a negative correlation with age (r =  −0.339; p < 0.001), maximum IOP (r =  −0.127; p = 0.006), and the interval since symptom onset (r =  −0.172; p < 0.001). The ECD was lower in eyes with ocular hypertension compared to those without ocular hypertension (p < 0.001). Eyes with KPs distributed on the central corneal endothelium had a significantly lower ECD than those with KPs distributed diffusely or KPs distributed triangularly on the inferior corneal endothelium (p = 0.006).Our findings suggest decreased ECD, increased cell size, and morphological alterations in the affected eyes of Chinese patients with FUS. The reduction in ECD is correlated with age, elevated IOP, the interval since symptom onset, and the distribution of KPs. [ABSTRACT FROM AUTHOR]

The corneal endothelium is responsible for pumping fluid out of the stroma in order to maintain corneal transparency, which depends in part on the expression and activity of sodium–potassium pumps. In this study, we evaluated how physiologic pressure and flow influence transcription, protein expression, and activity of Na+/K+-ATPase. Native and engineered corneal endothelia were cultured in a bioreactor in the presence of pressure and flow (hydrodynamic culture condition) or in a Petri dish (static culture condition). Transcription of ATP1A1 was assessed using qPCR, the expression of the α1 subunit of Na+/K+-ATPase was measured using Western blots and ELISA assays, and Na+/K+-ATPase activity was evaluated using an ATPase assay in the presence of ouabain. Results show that physiologic pressure and flow increase the transcription and the protein expression of Na+/K+-ATPase α1 in engineered corneal endothelia, while they remain stable in native corneal endothelia. Interestingly, the activity of Na+/K+-ATPase was increased in the presence of physiologic pressure and flow in both native and engineered corneal endothelia. These findings highlight the role of the in vivo environment on the functionality of the corneal endothelium. [ABSTRACT FROM AUTHOR]

Fuchs endothelial corneal dystrophy is a heterogenous disease with multifactorial etiology, and genetic, epigenetic, and exogenous factors contributing to its pathogenesis. DNA damage plays a significant role, with ultraviolet-A (UV-A) emerging as a key contributing factor. We investigate the potential application of neuropeptide α-melanocyte stimulating hormone (α-MSH) in mitigating oxidative stress induced endothelial damage. First, we examined the effects of α-MSH on a cultured human corneal endothelial cell line (HCEnC-21T) exposed to hydrogen peroxide (H2O2) induced oxidative DNA damage. We performed immunofluorescence and flow cytometry to assess DNA damage and cell death in the cultured cells. Additionally, we used an established mouse model that utilizes ultraviolet light to induce corneal endothelial cell damage resulting in decreased CEnC number, increased cell size variability, and decreased percentage of hexagonal cells. This endothelial decompensation leads to an increase in corneal thickness. Following UV-A exposure, the mice were systemically treated with α-MSH, either immediately after exposure (early treatment) or beginning two weeks post-exposure (delayed treatment). To evaluate treatment efficacy, we analyzed CEnC density and morphology using in vivo confocal microscopy, and central corneal thickness using anterior segment optical coherence tomography. Our findings demonstrated that α-MSH treatment effectively protects HCEnC-21T from free-radical induced oxidative DNA damage and subsequent cell death. In vivo, α-MSH treatment, mitigated the loss of CEnC density, deterioration of cell morphology and suppression of the resultant corneal swelling. These results underline the potential application of α-MSH as a therapeutic agent for mitigating corneal endothelial damage. [ABSTRACT FROM AUTHOR]

Descemet's Stripping Only (DSO) is a surgical technique that utilizes the peripheral corneal endothelial cell (CEnC) migration for wound closure. Ripasudil, a Rho-associated protein kinase (ROCK) inhibitor, has shown potential in DSO treatment; however, its mechanism in promoting CEnC migration remains unclear. We observed that ripasudil-treated immortalized normal and Fuchs endothelial corneal dystrophy (FECD) cells exhibited significantly enhanced migration and wound healing, particularly effective in FECD cells. Ripasudil upregulated mRNA expression of Snail Family Transcriptional Repressor (SNAI1/2) and Vimentin (VIM) while decreasing Cadherin (CDH1), indicating endothelial-to-mesenchymal transition (EMT) activation. Ripasudil activated Rac1, driving the actin-related protein complex (ARPC2) to the leading edge, facilitating enhanced migration. Ex vivo studies on cadaveric and FECD Descemet's membrane (DM) showed increased migration and proliferation of CEnCs after ripasudil treatment. An ex vivo DSO model demonstrated enhanced migration from the DM to the stroma with ripasudil. Coating small incision lenticule extraction (SMILE) tissues with an FNC coating mix and treating the cells in conjunction with ripasudil further improved migration and resulted in a monolayer formation, as detected by the ZO-1 junctional marker, thereby leading to the reduction in EMT. In conclusion, ripasudil effectively enhanced cellular migration, particularly in a novel ex vivo DSO model, when the stromal microenvironment was modulated. This suggests ripasudil as a promising adjuvant for DSO treatment, highlighting its potential clinical significance. [ABSTRACT FROM AUTHOR]

The experimental study of preservation length on the effect of high potassium concentration in the medium on the volume of human corneal endothelial cells was done. The results of the study of individual samples of fragments of donor material and the values calculated using joined data after hypothermic preservation for 4 and 10 days are presented. The increase of the time when cornea samples are kept in preservation medium (Eusol-C) at 4°C led to a decrease in the average value (M ± SEM) of cell swelling indicator (Q) in a potassium medium from 1.055 ± 0.001; n = 982 to 1.014 ± 0.001; n = 338; after 4 and 10 days, respectively. Student's t-test for independent samples showed a high degree of significance for the difference between these values (p = 2E-76). Identification of the proportion of cells capable to swell in a medium with a high content of potassium ions (Q ≥ 1), reflecting the electrogenic activity in these cells, showed a decrease of this indicator in the studied groups with increasing duration of preservation (94.3% and 56.8% after 4 and 10 days, respectively). Based on the results of the study, it is suggested that the values of endothelial cells swelling in a potassium environment can serve as indicators of the cells' ability to restore electrogenic transport. It is concluded that the study of cell swelling in a medium with a high content of potassium ions can provide information for predicting the functionality of the graft. [ABSTRACT FROM AUTHOR]

Purpose: To determine if smoking has an effect on corneal endothelium. Study Design: Descriptive observational. Place and Duration of Study: Lamar eye center in Kerbala city in Iraq during December, 2023 and January, 2024. Methods: A total of 108 eyes were selected, 54 eyes of smokers and 54 eyes of non-smokers. The smokers included in this study had history of smoking for more than twenty years and they smoke more than 40 cigarettes per day. All the participants were male. Specular microscopy was used to determine four corneal parameters; cell density, hexagonality, coefficient of variation and central corneal thickness. Results: Patients age ranged from 50 to 60 years. Central Corneal Thickness was 515.29±40.62 and 522.37±36.68 in smokers and non-smokers respectively with p=0.32. Coefficient of variation and, Cell Density and Hexagonal shape parameters were also not statistically different between the two groups (p>0.05). Conclusion: Smoking does not have a statistically significant effect on the corneal endothelium in terms of cell density, hexagonality, coefficient of variation, and central corneal thickness. However, more studies are needed to provide further evidence. [ABSTRACT FROM AUTHOR]

Introduction: Corneal graft detachment is a major postoperative complication of Descemet’s stripping automated endothelial keratoplasty (DSAEK). When a corneal graft becomes detached, corneal endothelial function generally fails, and repeat corneal transplantation is required. Herein, we report a rare case in which a transparent cornea was maintained after the removal of a dislocated DSAEK graft. Case Presentation: A 79-year-old woman with a residual lens cortex who had undergone cataract surgery was referred to our hospital. The cortex was removed, and bullous keratopathy progressed. Six months after the initial surgery, DSAEK was performed under topical anesthesia without any complications. Although the corneal graft had attached fairly well, it detached from the host cornea 3 weeks later. Two months after DSAEK, an air tamponade was used to treat the anterior chamber with single interrupted suturing; however, the graft detached again, except for the suture site. Because the detached cornea became cloudy in the anterior chamber, it was surgically removed 8 months after DSAEK. Accordingly, the host cornea transparency improved to a best-corrected visual acuity of 0.8 with a rigid gas permeable lens and a central corneal thickness of 580 μm. The corneal endothelial cell density was 995 cells/mm2. Conclusion: Removal of the corneal graft from the dislocated cloudy graft improved the visual acuity of this patient after DSAEK. The condition of the cornea should be carefully monitored after corneal endothelial transplantation, even after the graft has been dislocated. [ABSTRACT FROM AUTHOR]

Background: The aim of this review was to investigate the influence of various laser refractive surgery methods on the corneal endothelium in myopic patients. The role of the corneal endothelium in laser refractive surgery (LRS) is currently being addressed in the assessment of postoperative corneal edema risk. Methods: Changes in corneal endothelial cell density and morphology after LRS were evaluated based on a systematic review of current studies. The results of a literature search in the PubMed, Science Direct, Google Scholar, and the Web of Science databases, as well as a manual search, were selected for the final review according to the PRISMA 2020 flow diagram. Results: We included 24 prospective clinical trials in the review: surface ablation (twelve), LASIK and FemtoLASIK (two), femtosecond lenticule extraction (two), and comparable studies (eight). Endothelial cell density was determined by specular or in vivo confocal microscopy. In most studies, no statistically significant differences were found between preoperative and postoperative endothelial parameters. In nine studies, the changes were statistically significant, but no vision-threatening complications occurred, and no serious corneal complications developed in any eyes during the follow-up period. Conclusions: Based on collected data, laser keratorefractive surgery appears not to exert a significant effect on the corneal endothelium. [ABSTRACT FROM AUTHOR]

Background: Traumatic injuries in eyes previously treated with Deep Anterior Lamellar Keratoplasty (DALK) can lead to ruptures in the Descemet Membrane (DM) and damage to the corneal endothelium, a crucial layer for maintaining corneal clarity. Due to cell cycle constraints, the human corneal endothelium cannot proliferate; instead, it compensates for injury through cell enlargement and migration from adjacent areas. Methods: This study examines a notable case of corneal endothelial cell migration following a penetrating eye injury in a patient previously treated with DALK for keratoconus, supplemented by a review of relevant literature to contextualize the regenerative response. Results: A 39-year-old male with a history of DALK suffered a traumatic eye injury, resulting in damage to the Descemet Membrane and loss of the crystalline lens. After primary repair and considerations for further surgery, the patient's cornea cleared remarkably, with an improved visual acuity. This demonstrates the DM's potential for self-repair through endothelial cell migration. Conclusions: The outcomes suggest that delaying corneal transplant surgery for up to 3 months following Descemet Membrane injury due to ocular trauma could be advantageous. Allowing time for natural healing processes might eliminate the need for further invasive surgeries, thereby improving patient recovery outcomes. [ABSTRACT FROM AUTHOR]

Background: The aim of this study was to compare early clinical results regarding the safety and efficacy of dual-linear vs. conventional foot-pedal control in cataract surgery. Methods: This was a paired-eye contralateral, retrospective, observational study. Each patient underwent cataract surgery in both eyes: one eye with dual-linear foot-pedal control (study group) and the other eye with conventional foot-pedal control (control group). Absolute phaco time (APT), average phaco power, effective phaco time (EPT), and surgical complications were analyzed and compared. Corneal endothelial cell count, corneal thickness, corneal volume, and best-corrected distance visual acuity (BCDVA) were measured preoperatively and at 1 week, 1 month, and 3 months postoperatively. Results: A total of 94 patients (188 eyes) were enrolled. The respective APT, average phaco power, and EPT values were 7.05 ± 5.31 s, 28.4 ± 1.00, and 2.05 ± 1.56 s in the study group and 6.82 ± 6.48 s, 18.9 ± 1.74, and 1.35 ± 1.35 s in the control group. Conclusions: The average phaco power and EPT values were significantly higher in the study group. The safety of the dual-linear foot pedal was comparable to that of a conventional pedal in terms of endothelial cell loss, central corneal thickness, and surgical complications. [ABSTRACT FROM AUTHOR]

Purpose: To evaluate the effects of torsional and transverse phacoemulsification modes on corneal endothelial health. Material and Methods: Forty-nine eyes of 49 patients who were operated for cataracts were randomized into two groups. 27 patients in the first group were operated in the torsional phacoemulsification mode, and 22 patients in the second group were operated using the transversal mode. Preoperative, postoperative first day and postoperative first month; endothelial cell density (ECD), coefficient of variation (CV), hexagonality (HEX) and central corneal thickness (CCT) were analyzed. Results: The mean age was 67.89 ± 9.98 years in the first group and 66.77 ± 10.78 years in the second group. There was no significant difference between the groups in terms of mean age and ultrasound power (p=0.71 and p=0.26). While CCT increased significantly on the postoperative first day in both groups (p=0.001 and p=0.002), it returned to baseline at the postoperative first month. Although ECD decreased significantly at postoperative first month in both study groups (p=0.001 and p=0.005), it was above 1400 cells/µ2 in all cases. When the ECD, CV, 6A and CCT measurements performed before the operation, on the postoperative first day and at the postoperative first month were compared between the groups, it was observed that there was no significant difference. Conclusion: It was evaluated that phaco surgery performed with both modes is safe in terms of corneal thickness and endothelial health. However, it should be considered that phaco surgery slightly reduces the endothelial number and endothelial health should be evaluated before surgery. [ABSTRACT FROM AUTHOR]

Background Loss of endothelial cells is inevitable after cataract surgery. Therefore, it is critical to protect the corneal endothelium during cataract surgery to reach good visual outcomes afterward. Maintenance of corneal clarity depends on the good function of healthy endothelial cells in addition to an intact pumping system intended to remove fluid from the corneal stoma. Purpose Our study aimed to assess changes in the corneal endothelium using specular microscopy, after intracameral injection of ceftazidime in uncomplicated phacoemulsification and compared with a placebo. The changes were measured preoperatively and 1 month postoperatively. Patients and methods Our prospective cohort study comprised 40 eyes from 35 patients diagnosed with cataract, all of whom underwent uncomplicated phacoemulsification for cataract extraction. Results Endothelial cell density significantly decreased postoperatively compared with preoperative values in both groups. Central thickness of the cornea, coefficient of variation, and visual acuity significantly increased postoperatively compared to preoperative values in both groups. Endothelial cell density showed a significant decrease postoperatively with a P value equal to 0.01 for group A (placebo group) and 0.02 for group B (ceftazidime group). Central thickness of the cornea, coefficient of variation, and visual acuity showed significant increases postoperatively with a P value equal to 0.026, 0.005, and 0.001 for group A and 0.001, 0.034, and 0.001 for group B, respectively. Conclusion Ceftazidime can be considered safe for corneal endothelium when used intracamerally during phacoemulsification cataract surgery for prophylaxis of postoperative endophthalmitis. [ABSTRACT FROM AUTHOR]

On February 24–27, 2021, the Association for Ocular Pharmacology and Therapeutics (AOPT) held its 15th biennial scientific meeting online. The meeting was organized by Dr. Sanjoy Bhattacharya of the University of Miami in conjunction with the board of trustees of the AOPT. The 3-day conference was attended by academic scientists, clinicians, and industry and regulatory professionals. The theme of the meeting was Restoring Vision through Regeneration and it was sponsored, in part, by the National Institutes of Health, Bright Focus, Regeneron, and Santen (USA). During the 3 days of the meeting, presentations from several sessions explored different aspects of regenerative medicine in ophthalmology, including optic nerve regeneration, drugs and devices in glaucoma, retinal neuroprotection and plasticity, visual perception, and degeneration of trabecular meshwork. This article summarizes the proceedings of the session on corneal regenerative medicine research and discusses emerging concepts in drug development for corneal epithelial and endothelial regeneration. Since the meeting in 2021, several of these concepts have advanced to clinical-stage therapies, but so far as of 2023, none has been approved by regional regulatory authorities in the United States. One form of corneal endothelial cell therapy has been approved in Japan and only for bullous keratopathy. Ongoing work is proceeding in the United States and other countries. Clinical Registration No: National Clinical Trials 04894110, 04812667; Japan Registry for Clinical Trials a031210199. [ABSTRACT FROM AUTHOR]

Purpose: To investigate the corneal endothelial damage caused by acute primary angle closure (APAC) and related risk factors for severe corneal endothelial cell damage in Chinese subjects. Methods: In this multicentre retrospective study, 160 Chinese patients (171 eyes) diagnosed with APAC were recruited. Endothelial cell density (ECD) and morphological changes short after APAC were studied. Univariate regression and multivariate regression were used to identify risk factors associated with the extent of ECD reduction, including age, gender, education level, patients' location, systemic diseases, APAC duration (hours), highest recorded intraocular pressure (IOP), and presenting IOP. Factors associated with the probability of severe corneal damage (ECD lower than 1000/mm2) were analysed based on a linear function. Results: After one APAC episode, 12.28% eyes had ECD lower than 1000/mm2, 30.41% had ECD between 1000 and 2000/mm2, and 57.31% had ECD more than 2000/mm2. Attack duration was the only factor associated with severe endothelial damage (p < 0.0001). If the attack were to be subsided within 15.0 h, possibility of ECD lower than 1000/mm2 could be controlled under 1%. Conclusion: Shortly after the abortion of APAC, 12.28% patients experienced severe endothelial cell damage with ECD less than 1000/mm2. The only factor associated with severe ECD decrease was attack duration. Immediate and effective treatment is pivotal for preserving corneal endothelial function in APAC patients. [ABSTRACT FROM AUTHOR]

Purpose: To evaluate scleral thickness using anterior segment-optical coherence tomography (AS-OCT) in Fuchs endothelial dystrophy (FED) and compare the results with healthy individuals. Methods: Thirty-two eyes of 32 patients with FED and 30 eyes of 30 age, gender, spherical equivalent and axial length matched healthy participants were included. All subjects underwent a detailed ophthalmological examination including endothelial cell density and central corneal thickness (CCT) measurements. Scleral thickness was measured by AS-OCT (Swept Source-OCT, Triton,Topcon,Japan) in 4 quadrants (superior, inferior, nasal, temporal) from 6 mm posterior to the scleral spur. Results: The mean ages were 62.5 ± 13.2 (33–81) for FED group; 64 ± 8.1 (48–81) for control group. CCT was significantly greater in FED group than in the control group (586.8 ± 33.1 (514–635) vs 545.0 ± 20.7 (503–587), respectively)(p = 0.000). In FED group, mean scleral thickness in the superior, inferior, nasal and temporal quadrants were 434.0 ± 30.6 (371–498), 442.8 ± 27.6 (395–502), 447.7 ± 31.4 (382–502), 443.4 ± 30.3 (386–504) μm, respectively. In control group, the mean scleral thickness in the superior, inferior, nasal and temporal quadrants were 381.3 ± 20.0 (341–436), 383.2 ± 16.0 (352–436), 389.2 ± 21.0 (353–440), 383.2 ± 19.2 (349–440) µm, respectively. The mean scleral thickness was significantly higher in all quadrants in FED group than in control group (p = 0.000). Conclusion: In patients with FED, scleral thickness was significantly higher. FED is a progressive corneal disease that results in the accumulation of extracellular material in the cornea. These findings suggest that the accumulation of extracellular deposits may not be limited to the cornea. Due to their functional similarity and anatomical proximity, sclera may also be affected in FED. [ABSTRACT FROM AUTHOR]

Background: The quality of the endothelial cell layer is a major criterion for the approval of organ-cultured human donor-corneas for transplantation. We wanted to compare the predictive capacities of initial endothelial density and endothelium cell morphology for the approval of donor corneas for transplantation and for the clinical outcome after transplantation. Methods: The endothelial density and endothelium morphology in organ culture were examined by semiautomatic assessment of 1031 donor corneas. We performed a statistical analysis for correlations of donor-data and cultivation parameters regarding their predictive capacities for the final approval of donor corneas for transplantation and the clinical outcome of 202 transplanted patients. Results: Corneal endothelium cell density proved to be the only parameter with a certain predictive capacity with regard to the final decision, whether donor corneas are suitable for transplantation — however, the correlation was low (area under the curve [AUC] = 0.655). Endothelial cell morphology lacked any predictive power (AUC = 0.597). The clinical outcome regarding visual acuity seemed to be largely independent from both corneal endothelial cell density and morphology. Sub-analyses on transplanted patients stratified for their diagnoses vindicated these findings. Conclusions: Higher endothelial density (above a cut-off level of 2000 cells/mm2), as well as better endothelial morphology do not seem to be critical for transplant-corneal functionality in organ culture and up to 2 years after transplantation. Comparable long-term studies on graft survival are recommended to determine, whether the present endothelial density cut-off levels might be too stringent. [ABSTRACT FROM AUTHOR]

Rho kinase (ROCK) inhibitors have emerged as a key therapeutic class of interest in ophthalmology over the last decade. Promising in vitro studies laid the foundations for the development of novel therapeutic agents that target the ROCK signalling pathway in ocular disease, with subsequent clinical trials supporting their use. Corneal endothelial disease, glaucoma, and vitreoretinal disease are the major pathologies in which ROCK inhibitors have been investigated to date. Ripasudil and netarsudil represent the current leaders in this pharmaceutical group, having been extensively validated and approved for use in glaucoma in some countries. Less substantial evidence exists for fasudil in ophthalmic use. ROCK inhibitors are also increasingly used in cultured endothelial cell grafting and as an adjunct to aid in endothelial cell migration and replication in Descemet's stripping procedures or Descemet's membrane injuries. This review has synthesised both established and emerging research to provide a practical guide to prescribing in this drug class. Drug efficacies, side effect profiles, and the demographic and clinical characteristics of appropriate drug candidates are discussed. [ABSTRACT FROM AUTHOR]

Nowadays, treating corneal diseases arising from injury to the corneal endothelium necessitates donor tissue, but these corneas are extremely scarce. As a result, researchers are dedicating significant efforts to exploring alternative approaches that do not rely on donor tissues. Among these, creating a tissue-engineered scaffold on which corneal endothelial cells can be transplanted holds particular fascination. Numerous functional materials, encompassing natural, semi-synthetic, and synthetic polymers, have already been studied in this regard. In this review, we present a comprehensive overview of recent advancements in using polymer biomaterials as scaffolds for corneal endothelium tissue engineering. Initially, we analyze and present the key properties necessary for an effective corneal endothelial implant utilizing polymer biomaterials. Subsequently, we focus on various emerging biomaterials as scaffolds for corneal endothelium tissue engineering. We discuss their modifications (including natural and synthetic composites) and analyze the effect of micro- and nano-topological morphology on corneal endothelial scaffolds. Lastly, we highlight the challenges and prospects of these materials in corneal endothelium tissue engineering. [ABSTRACT FROM AUTHOR]

Purpose: To compare endothelial cell density (ECD), percentage of hexagonal cells (%Hex) and coefficient of variation (CV) in cell size following lens cataract surgery with phacoemulsification performed using Continuous Curvilinear Capsulorhexis (CCC) or Precision Pulse Capsulotomy (PPC). Patients and Methods: Sixty-seven subjects were randomly assigned to undergo lens cataract removal with the capsulotomy step performed using either CCC or PPC. Specular microscopy images were obtained pre-operatively, 1 month and 3 months after surgery. ECD, %Hex and CV were analyzed in a masked fashion by an independent reading center. Results: The mean percentage ECD loss at 1 month was 11.5% in the CCC group and 12.3% in the PPC group (P = 0.818; t-test). At 3 months, the mean percentage ECD loss was 11.7% in the CCC group and 12.4% in the PPC group (P = 0.815; t-test). The mean %Hex at 1 month was 54.3% in the CCC group and 54.7% in the PPC group (P = 0.695; t-test). At 3 months, the mean %Hex was 56.2% in the CCC group and 54.7% in the PPC group (P = 0.278; t-test). The CV at 1 month was 34.4% in the CCC group and 34.3% in the PPC group (P = 0.927; t-test). At 3 months, the CV was 32.7% in the CCC group and 33.4% in the PPC group (P = 0.864; t-test). Conclusion: No differences in ECD loss, %Hex and CV were observed between patients who received CCC or PPC. PPC use during cataract surgery does not result in any increased endothelial cell loss beyond that normally associated with this surgery. [ABSTRACT FROM AUTHOR]

Corneal endothelium is a single cell layer that is mainly responsible for maintaining corneal clarity. Endothelial damage secondary to toxicity, stress, or genetic predisposition are common and in conjunction with the low regenerative ability of the cells, making their preservation critical for maintaining visual acuity. Patients with glaucoma, who are estimated to be close to 80 million worldwide, have a plethora of reasons for developing endothelial damage, being exposed to a spectrum that extends from various medical and surgical interventions to the disease itself. The wide spectrum of glaucoma pharmacotherapy that has been recently extended by addition of newer classes of medications has been the focus of extensive research on its effects on corneal endothelium. Both basic and clinical research have attempted to shine a light on the complex mechanisms associated with the effects of glaucoma medication on corneal endothelium and to answer the important question as to whether these findings are clinically significant. The aim of this review is to summarize and present current literature of the various findings, both from in vivo and in vitro studies that have focused on the complex relationship between different classes of glaucoma medication and their effect on corneal endothelium. [ABSTRACT FROM AUTHOR]

Purpose: In many epithelia, including the corneal endothelium, intracellular/extracellular stresses break down the perijunctional actomyosin ring (PAMR) and zonula occludens-1 (ZO-1) at the apical junctions. This study aims to grade the severity of damage to PAMR and ZO-1 through machine learning. Methods: Immunocytochemical images of PAMR and ZO-1 were drawn from recent studies on the corneal endothelium subjected to hypothermia and oxidative stress. The images were analyzed for their morphological (e.g., Hu moments) and textural features (based on gray-level co-occurrence matrix [GLCM] and Gabor filters). The extracted features were ranked by SHapley analysis and analysis of variance. Then top features were used to grade the severity of damage using a suite of ensemble classifiers, including random forest, bagging classifier (BC), AdaBoost, extreme gradient boosting, and stacking classifier. Results: A partial set of features from GLCM, along with Hu moments and the number of hexagons, enabled the classification of damage to PAMR into Control, Mild, Moderate, and Severe with the area under the receiver operating characteristics curve (AUC) = 0.92 and F1 score = 0.77 with BC. In contrast, a bank of Gabor filters provided a partial set of features that could be combined with Hu moments, branch length, and sharpness for the classification of ZO-1 images into four levels with AUC = 0.95 and F1 score of 0.8 with BC. Conclusions: We have developed a workflow that enables the stratification of damage to PAMR and ZO-1. The approach can be applied to similar data during drug discovery or pathophysiological studies of epithelia. [ABSTRACT FROM AUTHOR]

(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]

(1) Background: Cell injection therapy is an emerging treatment for bullous keratopathy (BK). Anterior segment optical coherence tomography (AS-OCT) imaging allows the high-resolution assessment of the anterior chamber. Our study aimed to investigate the predictive value of the visibility of cellular aggregates for corneal deturgescence in an animal model of bullous keratopathy. (2) Methods: Cell injections of corneal endothelial cells were performed in 45 eyes in a rabbit model of BK. AS-OCT imaging and central corneal thickness (CCT) measurement were performed at baseline and on day 1, day 4, day 7 and day 14 following cell injection. A logistic regression was modelled to predict successful corneal deturgescence and its failure with cell aggregate visibility and CCT. Receiver-operating characteristic (ROC) curves were plotted, and areas under the curve (AUC) calculated for each time point in these models. (3) Results: Cellular aggregates were identified on days 1, 4, 7 and 14 in 86.7%, 39.5%, 20.0% and 4.4% of eyes, respectively. The positive predictive value of cellular aggregate visibility for successful corneal deturgescence was 71.8%, 64.7%, 66.7% and 100.0% at each time point, respectively. Using logistic regression modelling, the visibility of cellular aggregates on day 1 appeared to increase the likelihood of successful corneal deturgescence, but this did not reach statistical significance. An increase in pachymetry, however, resulted in a small but statistically significant decreased likelihood of success, with an odds ratio of 0.996 for days 1 (95% CI 0.993–1.000), 2 (95% CI 0.993–0.999) and 14 (95% CI 0.994–0.998) and an odds ratio of 0.994 (95% CI 0.991–0.998) for day 7. The ROC curves were plotted, and the AUC values were 0.72 (95% CI 0.55–0.89), 0.80 (95% CI 0. 62–0.98), 0.86 (95% CI 0.71–1.00) and 0.90 (95% CI 0.80–0.99) for days 1, 4, 7 and 14, respectively. (4) Conclusions: Logistic regression modelling of cell aggregate visibility and CCT was predictive of successful corneal endothelial cell injection therapy. [ABSTRACT FROM AUTHOR]

Background: Pseudoexfoliation causes characteristic corneal endothelial changes leading to corneal decompensation and loss of corneal transparency severely affecting vision. Objectives: To evaluate morphological changes in corneal endothelial cells and central corneal thickness in pseudoexfoliation syndrome (PXS) and to compare it with age matched patients without pseudoexfoliation syndrome. Material and Methods: A hospital based, cross sectional analytical study was conducted at Upgraded Department of Ophthalmology, Government Medical College/hospital, Jammu in which total of 46 patients planned for cataract surgery were included (23 patients with pseudoexfoliation syndrome and 23 patients without pseudoexfoliation syndrome). Corneal endothelial cell density (ECD), percentage of hexagonal cells, coefficient of variation (CV) in cell size, and central corneal thickness (CCT) were measured using specular microscopy. Results: The mean ECD was 2325.1 ± 383.1 in the PXS group and 2763 ± 351.3 in the control group respectively, and ECD in the PXS group was significantly lower than in that in the control group (p=0.0002). The percentage of hexagonal cells was 48.3 ± 5.7 in PXS group and 51.5 ± 4.6 in the control group respectively, and percentage of hexagonal cells was significantly lower than that in the control group (p=0.04). The cell size coefficient of variation was 33.9 ± 2.6 in the PXS group and 33.0 ± 3.1 in the control group respectively, and patients with PXS had no statistically significant difference in cell size coefficient of variation compared to control group (p=0.29). The CCT was 509.0 ± 27.3 in the PXS group and 529.0 ± 23.6 in the control group respectively, and CCT in the PXS group was significantly lower than that in the control group (p=0.01). Conclusion: In our study the ECD and CCT was significantly lower in patients with pseudoexfoliation syndrome regardless of the presence of pseudoexfoliative glaucoma. [ABSTRACT FROM AUTHOR]

The aim of this study was to evaluate the influence of phacoemulsification cataract surgery on the state of the corneal endothelium in diabetic versus non-diabetic patients. We compared the corneal cell morphology in 48 diabetics with good glycemic control and 72 non-diabetic patients before and after uneventful phacoemulsification. Corneal cell density, central corneal thickness, and hexagonality were measured preoperatively and post-surgery (at 1 and 4 weeks) by specular microscopy. The effect of age, gender, axial length, and anterior chamber depth on the parameters of the corneal endothelium were evaluated. We noticed a drop in the endothelial density in both groups postoperatively: a mean endothelial cell loss of 472.7 ± 369.1 in the diabetic group was recorded versus 165.7 ± 214.6 mean loss in the non-diabetic group after the first week. A significant increase in central corneal thickness was also noticed in both groups one week after phacoemulsification, but no statistical significance after 4 weeks in the diabetic group. In terms of cell hexagonality, statistically significant differences were noticed after 4 weeks in both groups. Overall, a significant difference between diabetic and non-diabetic population was noticed in terms of corneal endothelial cell loss after uneventful phacoemulsification cataract surgery. Routine specular microscopy and HbA1c evaluation is recommended before cataract surgery, while intraoperative precautions and high monitorisation in terms of pacho power intensity and ultrasound energy, along with a proper application of the dispersive viscoelastic substances are essential to reduce the risk of endothelial damage. [ABSTRACT FROM AUTHOR]

To demonstrate the suitability of using decellularized SMILE (Small-incision Lenticule Extraction) lenticules for culturing and transplanting the corneal endothelium (CE). The SMILE lenticules, obtained during refractive surgery, were decellularized by incubating in CE culture medium and fetal bovine serum. Decellularization was confirmed by hematoxylin and eosin staining, DAPI staining, and gel electrophoresis. The amount of DNA per milligram of dry tissue weight was calculated to quantify the residual nuclear content. The transparency of the decellularized lenticules was determined by calculating the modulation transfer function. Immunostaining for stromal collagens and glycosaminoglycan was performed using specific antibodies. Engineered tissue was constructed by culturing the CE cells on lenticules and staining for ZO-1, Na/K ATPase, and N-cadherin. The functionality of the engineered tissues was assessed by transplanting them onto edematous human donor corneas and perfusing for 10 days ex-vivo. The residual DNA per milligram of dry tissue weight was found to be significantly reduced (p < 0.0001) in serum (0.255 µg/mg) and Opti-MEM (0.140 µg/mg) when compared to fresh lenticules (3.9 µg/mg). Decellularization did not alter the arrangement of the collagen fibers or the transparency of the lenticules. CE cells attached and matured to express ZO-1, Na/K ATPase, and N-cadherin at two weeks after seeding. The engineered tissue upon transplantation significantly reduced the corneal edema (p < 0.05) and the transplanted cells remained intact on the SMILE lenticule post-transplantation. This study demonstrates the suitability of using SMILE lenticules decellularized using a simple, chemical-free method for engineering the corneal endothelium for transplantation. [ABSTRACT FROM AUTHOR]

Background: phacoemulsification is the preferred surgical procedure in treatment of cataract nowadays. Its outcome can be affected by health of the cornea, intra ocular pressure, manipulation and time of surgery in mature and white cataracts, trypan blue staining of the anterior capsule improves capsule visibility. Aim: To assess Trypan blue stain's effectiveness and safety after cataract surgery. Patients and Methods: This prospective cohort study's was performed at Ophthalmology Department, faculty of Medicine, Zagazig University hospitals. Patients with cataract (nuclear II III) or PSC, age group from (20-60) and patients with more than 100 corneal endothelial cells per 2000 cells/mm2 were covered by the research. They were divided into the case group and control group. All patients were subjected to history taking, complete slit-lamp examination including grading the density of the cataract and an indirect ophthalmoscopy examination. Results: There was significant decrease in ECD in both groups after one month of phaco surgery with statistically highly significant decrease in group 2 compared to group 1. The same was observed in hexagonality and coefficient of variance. Central corneal thickness showed no difference in change between both groups. Conclusion: There aren't significant alterations in the corneal endothelium one month postoperatively after staining anterior capsule by trypan blue 1% in phacoemulsification surgery. [ABSTRACT FROM AUTHOR]

Here, we report the case of a patient with multiple parallel-line endotheliitis with myopic shift, which has not been previously reported. A 36-year-old man visited our clinic with blurred vision in his left eye. A slit lamp examination revealed an interesting pattern of multiple parallel lines of keratic precipitates, along with subtle corneal edema. The spherical equivalents measured in the right and left eyes were −9.25 and −11.875 diopter, respectively, with the left eye showing more myopic progression. On specular microscopy, endothelial cell loss was found in the left eye. After administration of a weak topical steroid, keratic precipitates and corneal edema resolved completely within 2 days. The myopic shift in his left eye recovered to −10.0 diopters. Multiple parallel-line endotheliitis can be easily treated with a weak topical steroid in a short period. A temporary myopic shift may occur, which in this case resolved upon disease recovery. However, corneal endothelial cell loss is inevitable; therefore, careful monitoring is needed. [ABSTRACT FROM AUTHOR]

Purpose: The fluid pump and barrier functions of the corneal endothelium maintain stromal deturgescence required for corneal transparency. The effect of oxidative stress, a hallmark of Fuchs endothelial corneal dystrophy (FECD), on the endothelial barrier function has been investigated. Methods: The endothelium of porcine corneas ex vivo was exposed to (1) membrane permeable oxidants (H2O2, 100 μM, 1 h; tert-butyl-hydroperoxide, 100 μM, 1 h), or (2) ultraviolet A (UVA) with photosensitizers for 15 min, riboflavin (50 μM) or tryptophan (Trp) (100 μM). The effects on the apical junction complex were analyzed by (1) immunostaining the perijunctional actomyosin ring (PAMR) and ZO-1 and (2) assessment of paracellular flux of fluorescein isothiocyanate (FITC)-avidin across cultured endothelial cells grown on biotinylated-gelatin film. The extent of oxidative stress was quantified by changes in intracellular reactive oxygen species (ROS) and mitochondrial membrane potential (MMP) in addition to lipid peroxidation and release of lactate dehydrogenase (LDH). Results: Both methods of oxidative stress led to the disruption of PAMR and ZO-1 concurrent with changes in ROS levels, depolarization of MMP, increased lipid peroxidation, elevated LDH release, and increased permeability of FITC-avidin. The effects of direct oxidants were opposed by SB-203580 [p38 mitogen-activating protein (MAP) kinase inhibitor; 10 μM]. The damage by UVA+photosensitizers was blocked by extracellular catalase (10,000 U/mL). Conclusions: (1) Acute oxidative stress breaks down the barrier function through destruction of PAMR in a p38 MAP kinase-dependent manner. (2) UVA+photosensitizers elicit the breakdown of PAMR via type I reactions, involving H2O2 release. (3) Blocking the oxidative stress prevents loss of barrier function, which could be helpful in the therapeutics of FECD. [ABSTRACT FROM AUTHOR]

Purpose: The most crucial step in deep anterior lamellar keratoplasty (DALK) is to achieve a bare Descemet's membrane. We aimed to assess a new femtosecond laser software that allows for a precise intrastromal tunnel creation for big bubble (BB) air injection using a real-time microscope-integrated optical coherence tomography. Materials and methods: A retrospective review of 61 eyes of 61 patients with keratoconus. Before introducing the new software update, DALK was performed using a partial-assisted femtosecond laser (partial-thickness circular cut followed by a lamellar cut) with manual intrastromal tunnel creation (partial FS-DALK group). After the software update, the femtosecond laser created the intrastromal tunnel (full FS-DALK group). Results: In the full FS-DALK group, the BB's formation was significantly higher (64.3% vs. 36.4%, p = 0.04), and surgery time was shorter (21.8 ± 5.1 vs. 25.6 ± 6.8 min, p = 0.025) than in the partial FS-DALK. Penetrating keratoplasty conversion rate (7.1% vs. 15.1%, p = 0.432) was similar between the groups. Both groups showed statistically significant improvement in uncorrected and corrected distance visual acuity, central corneal thickness, surface asymmetry, and regularity indices. Endothelial cell density loss at 12 and 18 months was lower in the full compared with the partial FS-DALK group (12 months:10.0% vs. 16; 18 months: 10.7 vs. 16.5%, p < 0.001 for both comparisons). Conclusions: Creating the intrastromal guiding tunnel using FS laser for air injection resulted in a higher rate of BB formation, reduced long-term endothelial cell loss, and operating room time. [ABSTRACT FROM AUTHOR]

Dry eye disease (DED) and other inflammatory ocular disorders have been reported to be associated with decreased corneal endothelial cell density (CECD), however the mechanism of underlying endothelial cell loss remains unknown. We conducted a comprehensive literature search of English-written publications on dry eye disease, corneal endothelial cell loss, Sjögren's syndrome, and Graft Vs Host Disease (GVHD), to review the effects of DED and other inflammatory ocular surface conditions on CECD. A total of 78 studies were included in our study. Loss of corneal neurotrophic support, cytotoxic stress, and a heightened immune response, all of which may occur secondarily to a common causative agent such as inflammation, are major contributors to reduced CECD. More studies are needed to determine how the interrelated pathways of altered corneal nerve function and upregulated expression of inflammatory activity influence corneal endothelial cell loss. [ABSTRACT FROM AUTHOR]

Diabetic mellitus (DM) causes various complications, including the corneal endothelial dysfunction that leads to corneal edema and vision loss, especially in the DM patients with intraocular surgeries. However, the pathogenic mechanism of hyperglycemia-caused corneal endothelial dysfunction remains incomplete understood. Here we firstly screened and identified the glutamine contents of aqueous humor (AH) were significantly reduced in the type 2 diabetic patients and type 1 and type 2 diabetic mice. To explore the potential therapeutic effects of glutamine (Gln) supplement on the protection of diabetic corneal endothelial dysfunction, we performed the anterior chamber perfusion with the addition of L-alanyl-L-glutamine (Ala-Gln), and confirmed that Ala-Gln supplement not only accelerated the resolution of corneal edema and recovery of corneal thickness, but also preserved the regular arrangement and barrier-pump function of cornea. Mechanistically, we revealed that the supplements of Ala-Gln protect corneal endothelial cells (CECs) from the deleterious effects of high glucose-induced oxidative stress, mitochondrial dysfunction, and cell apoptosis. Overall, these results indicate the Gln depletion plays an important role in the diabetic corneal endothelial dysfunction, while the Ala-Gln supplement during intraocular surgery provide an effective prevention strategy through regulating the redox homeostasis and mitochondrial function of corneal endothelium. • The glutamine levels in aqueous humor were obviously decreased in diabetic patients, as well as in diabetic mice. • High glucose environment leads to oxidative stress and mitochondrial dysfunction in corneal endothelial cells. • Glutamine supplementation could promote the recovery of diabetic corneal endothelial cells after damage. [ABSTRACT FROM AUTHOR]

Background: Endothelial cell density (ECD), morphology and central corneal thickness (CCT) are important because they facilitate the assessment of the functional reserve of the endothelium in patients before any intraocular surgery. Aim of the Work: The goal of this study was to measure the changes in corneal endothelium using non-contact specular microscopy in eyes that underwent uneventful phacoemulsification and intraocular lens implantation. Patients and Methods: A randomized controlled clinical trial was conducted for a period of six months on a group of 50 patients at the Ophthalmology Department, Ain Shams University Hospital, half of which underwent phacoemulsification and IOL implantation with injection of intracameral epinephrine 1:10,000 with sodium bisulfite as a preservative, and the other half underwent the same procedure without injection of intracameral epinephrine. Results: There was no statistically significant difference between both groups regarding CD (Cell Density), with a P-value = 0.089, and also regarding SD (Standard Deviation), CV (Coefficient of Variation), HEX (Hexagonal Cells) and CCT (Central Corneal Thickness), with P-value 0.177, 0.770, 0.144 and 0.961 respectively. Conclusion: Intracameral epinephrine 1:10,000 with sodium bisulfite could improve pupillary dilatation and is safe on the corneal endothelium as well. [ABSTRACT FROM AUTHOR]

Fuchs endothelial corneal dystrophy (FECD), a degenerative corneal condition, is characterized by the droplet-like accumulation of the extracellular matrix, known as guttae and progressive loss of corneal endothelial cells ultimately leading to visual distortion and glare. FECD can be influenced by environmental stressors and genetic conditions. However, the role of mitochondrial dysfunction for advancing FECD pathogenesis is not yet fully studied. Therefore, in the present study we sought to determine whether a combination of environmental stressors (ultraviolet-A (UVA) light and cigarette smoke condensate (CSC)) can induce mitochondrial dysfunction leading to FECD. We also investigated if MitoQ, a water-soluble antioxidant, can target mitochondrial dysfunction induced by UVA and CSC in human corneal endothelial cells mitigating FECD pathogenesis. We modeled the FECD by increasing exogenous oxidative stress with CSC (0.2%), UVA (25J/cm2) and a combination of UVA + CSC and performed a temporal analysis of their cellular and mitochondrial effects on HCEnC-21T immortalized cells in vitro before and after MitoQ (0.05 μM) treatment. Interestingly, we observed that a combination of UVA + CSC exposure increased mitochondrial ROS and fragmentation leading to a lower mitochondrial membrane potential and increased levels of cytochrome c release leading to apoptosis and cell death. MitoQ intervention successfully mitigated these effects and restored cell viability. The UVA + CSC model could be used to study stress induced mitochondrial dysfunction. Additionally, MitoQ can serve as a viable antioxidant in attenuating mitochondrial dysfunction, underscoring its potential as a molecular-focused treatment approach to combat FECD pathogenesis. • The study elucidates the role of mitochondrial dysfunction in Fuchs endothelial corneal dystrophy (FECD) shedding light on a potential mechanism driving its pathogenesis. • Environmental stressors such as ultraviolet-A (UVA) light and cigarette smoke condensate (CSC) are identified as contributors to mitochondrial dysfunction, providing insight into the interplay between external factors and cellular health in FECD development. • MitoQ, a water-soluble antioxidant, emerges as a promising intervention to mitigate mitochondrial dysfunction induced by UVA and CSC exposure. Its ability to restore cell viability underscores its potential as a targeted treatment for FECD. • The combination of UVA and CSC exposure is proposed as a model to study stress-induced mitochondrial dysfunction relevant to FECD, offering researchers a valuable tool for further investigation into the condition's underlying mechanisms. [ABSTRACT FROM AUTHOR]

The endothelial layer of the cornea plays a critical role in regulating its hydration by actively controlling fluid intake in the tissue via transporting the excess fluid out to the aqueous humor. A damaged corneal endothelial layer leads to perturbations in tissue hydration and edema, which can impact corneal transparency and visual acuity. We utilized a non-contact terahertz (THz) scanner designed for imaging spherical targets to discriminate between ex vivo corneal samples with intact and damaged endothelial layers. To create varying grades of corneal edema, the intraocular pressures of the whole porcine eye globe samples (n = 19) were increased to either 25, 35 or 45 mmHg for 4 h before returning to normal pressure levels at 15 mmHg for the remaining 4 h. Changes in tissue hydration were assessed by differences in spectral slopes between 0.4 and 0.8 THz. Our results indicate that the THz response of the corneal samples can vary according to the differences in the endothelial cell density, as determined by SEM imaging. We show that this spectroscopic difference is statistically significant and can be used to assess the intactness of the endothelial layer. These results demonstrate that THz can noninvasively assess the corneal endothelium and provide valuable complimentary information for the study and diagnosis of corneal diseases that perturb the tissue hydration. [ABSTRACT FROM AUTHOR]

Purpose: To characterize the impact of corneal cold storage (CS) on the endothelial apical junctional complex (AJC). Methods: Porcine corneas were held in CS (4°C; 1–7 days) with Cornisol™ preservation medium supplemented with epothilone B (EpoB; microtubule stabilizer; 100 nM), SB-203580 (p38 mitogen-activated protein [MAP] kinase inhibitor; 20 μM), or antioxidants (quercetin, 100 μM; vitamin E, 1 mM; deferoxamine, an iron chelator, 10 mM). After CS termination, the damage to endothelial AJC was characterized by imaging perijunctional actomyosin ring (PAMR) and zonula occludens (ZO-1). The effects of EpoB and SB-203580 were characterized by imaging microtubules. The loss in the barrier function was assessed in cultured cells grown on biotin-coated gelatin by permeability to fluorescein isothiocyanate (FITC)-avidin. The accumulation of reactive oxygen species (ROS), altered mitochondrial membrane potential (MMP), lipid peroxidation, and lactate dehydrogenase (LDH) release were also determined in response to CS. Results: CS led to the loss of microtubules, destruction of PAMR, and breakdown of ZO-1 in the endothelium. The severity of damage increased when CS was prolonged. Although rewarming of the tissue increased the damage, the effect was marginal. CS also induced accumulation of ROS, alteration in MMP, lipid peroxidation, enhanced LDH release, and increased permeability to FITC-avidin. These changes were opposed by EpoB, SB-203580, and antioxidants. Conclusion: Corneal CS destroys AJC of the endothelium, leading to loss of its barrier function. The effects were surmounted by microtubule stabilization, p38 MAP kinase inhibition, and antioxidants. Thus, there is potential for reformulation of the preservation medium to maintain the health of the donor corneal endothelium before transplantation. [ABSTRACT FROM AUTHOR]

Objectives: To evaluate the effects of intracameral drugs and dyes on rat corneal endothelial apoptosis and cell morphology. Materials and Methods: The right eyes of 72 rats were injected intracamerally with 1% lidocaine, 0.01% adrenaline, triamcinolone acetonide (TA) 4 mg/mL, 1% trypan blue (TB), 0.5% indocyanine green (ICG), and fortified balanced salt solution as control. Corneal samples were taken 1 day and 1 week post-injection. Corneal endothelial apoptosis was assessed by the TUNEL technique, and the ratio of apoptotic cells in each group was compared with the control. Corneal endothelial cell morphology was evaluated in each specimen by transmission electron microscopy. Results: The mean apoptotic endothelial cell ratio was significantly higher at 1 day and 1 week after intracameral adrenaline injection when compared to controls (p=0.03 and 0.021, respectively). TB caused a significantly higher apoptotic cell ratio when compared to controls at 1 week after injection (p=0.043). Lidocaine caused a higher apoptotic cell ratio compared to TA and ICG at 1 week, although not statistically significant (p=0.058, 0.09, 0.69, respectively). In all experimental specimens, transmission electron microscopy showed morphological changes associated with apoptosis. Conclusion: This study showed that intracameral adrenaline, TB, and lidocaine injections may have toxic effects on corneal tissue, as indicated by ultrastructural and histopathological alterations. Therefore, these agents should be used with caution in intraocular surgery. [ABSTRACT FROM AUTHOR]

Highlights: This study is the first prospective clinical trial to evaluate the anterior corneal parameters in patients with acromegaly before and after transsphenoidal pituitary adenoma resection on the basis of several months of postoperative follow-up. An increase in CD and a decrease in AVG and CCTNSM were observed after pituitary adenoma resection. The mean CCT measured both with an ultrasonic pachymeter and with non-contact specular microscopy was significantly lower as compared to the preoperative values. IOP decreased statistically for the measured IOP air puff values. A downward trend was observed after treatment in the case of IOP GAT and IOPc, but statistical significance was not achieved. Statistically significant reduction in CH and CRF was found after the surgical treatment of acromegaly. (1) Purpose: This study aimed to investigate the changes in Reichert Ocular Response Analyzer (ORA) parameters, corneal endothelium parameters, central corneal thickness (CCT), and intraocular pressure (IOP) before and after the transsphenoidal resection of pituitary adenoma in patients with acromegaly. (2) Methods: This was a single-center, prospective, interventional study. Twenty patients with newly diagnosed acromegaly were examined before and 19 ± 9 months after transsphenoidal resection. The participants underwent a comprehensive ophthalmological examination including pneumatic IOP (IOP air puff), Goldmann applanation tonometry (IOP GAT), CCT measured using the iPac pachymeter (CCTUP), IOP value corrected for CCTUP using the Ehlers formula (IOPc) ORA measurements included corneal hysteresis (CH), corneal resistance factor (CRF), corneal-compensated IOP (IOPcc), and Goldmann-correlated IOP (IOPg). CCT from non-contact specular microscopy (CCTNSM), the number of endothelial cells (CD) per mm2, and average cell size (AVG) were determined with non-contact specular microscopy. (3) Results: A statistically significant decrease was observed in CCTUP (p = 0.007), and IOP air puff (p = 0.012) after surgery. Moreover, we noted a statistically significant increase in CD (p = 0.001), and a statistically significant decrease in AVG (p = 0.009) and CCTNSM (p = 0.004) after surgery. A statistically significant decrease was also observed in IOPg (p = 0.011), CH (p = 0.016), and CRF (p = 0.001) after surgery. The mean value of IOP GAT and IOPc was lower after the surgery. However, the difference was not statistically significant. (4) Conclusions: Our study revealed significant changes in biomechanics, corneal endothelium, CCT and IOP after pituitary adenoma resection in patients with acromegaly. It proves that the eye might be sensitive to long-term overexposure to growth hormone (GH) and insulin-like growth factor-1 (IGF-1). We suggest that disease activity be taken into consideration on ophthalmological examination. [ABSTRACT FROM AUTHOR]

Cataract causes bilateral blindness in 20 million people globally, the vast majority of whom live in developing countries. Manual small-incision cataract surgery (MSICS) has emerged as an efficient and economical alternative to phacoemulsification, giving comparable results in terms of final visual gain. One of the important determinants of postoperative visual gain is the status of the corneal endothelium. Multiple factors such as corneal distortion, irrigation solution turbulence, mechanical trauma by instruments, nuclear fragments, intraocular lens contact, and free oxygen radicals, all have been implicated in causing corneal damage during cataract surgery. MSICS with posterior chamber intraocular lens implantation has been reported to cause an endothelial cell loss of 15.83%, which is comparable with other modes of cataract surgery like extracapsular cataract extraction and phacoemulsification. Thorough preoperative assessment of endothelial status and taking necessary steps for endothelial protection during surgery can decrease the endothelial cell loss and overall burden of pseudophakic bullous keratopathy. In addition to surgical techniques, the type of irrigating solutions, ocular viscoelastic devices, intracameral dyes, and drugs all affect the endothelial cell status. This review presents a summary of available literature on the protection of endothelial cells during different steps of MSICS. This is especially relevant for developing countries where large-scale MSICS cataract surgeries are performed to decrease the cataract blindness burden. [ABSTRACT FROM AUTHOR]

Purpose Posterior corneal vesicles (PCVs) have clinical features that are similar to posterior polymorphous corneal dystrophy (PPCD). To help determine whether there is a shared genetic basis, we screened 38 individuals with PCVs for changes in the three genes identified as causative for PPCD. Methods: We prospectively recruited patients for this study. We examined all individuals clinically, with their first‐degree relatives when available. We used a combination of Sanger and exome sequencing to screen regulatory regions of OVOL2 and GRHL2, and the entire ZEB1 coding sequence. Results: The median age at examination was 37.5 years (range 4.7–84.0 years), 20 (53%) were male and in 19 (50%) the PCVs were unilateral. Most individuals were discharged to optometric review, but five had follow‐up for a median of 12 years (range 5–13 years) with no evidence of progression. In cases with unilateral PCVs, there was statistically significant evidence that the change in the affected eye was associated with a lower endothelial cell density (p = 0.0003), greater central corneal thickness (p = 0.0277) and a steeper mean keratometry (p = 0.0034), but not with a higher keratometric astigmatism or a reduced LogMAR visual acuity. First‐degree relatives of 13 individuals were available for examination, and in 3 (23%), PCVs were identified. No possibly pathogenic variants were identified in the PPCD‐associated genes screened. Conclusion: We found no evidence that PCVs share the same genetic background as PPCD. In contrast to PPCD, we confirm that PCVs is a mild, non‐progressive condition with no requirement for long‐term review. However, subsequent cataract surgery can lead to corneal oedema. [ABSTRACT FROM AUTHOR]

Introduction: To elucidate the specific functions of the primary cilia in corneal endothelial cells (CECs) by investigating the histological changes of corneal endothelium exposed at low temperature. Study design: Experimental study. Methods: This study involved corneas freshly obtained from Japanese white rabbits preserved in Optisol™-GS (Bausch & Lomb) corneal storage medium at 4 °C for 0, 1, and 7 days. Corneas preserved for 7 days were also incubated at 37 °C in culture media for an additional 2 days. A rabbit CEC line was also preserved in Optisol™-GS at 4 °C for 0 and 1 day. The corneal endothelium specimens and CECs were then assessed by immunostaining and scanning electron-microscopy (SEM). Results: Immediately post isolation, the CECs of the specimens showed positive immunostaining for primary cilia (i.e., approximately 20%) via anti-acetylated alpha Tubulin antibody and SEM observation. Primary cilia were found to have attenuated/disappeared on the corneal endothelium specimens preserved for 1 or 7 days at 4 °C. After an additional 2-day incubation at 37 °C, primary cilia reappeared on the corneal endothelium specimens (approximately 20%). The disappearance of cilia during the preservation period was also observed in the immortalized CECs. Conclusion: The findings in this study using rabbit corneas indicate that the primary cilia of corneal endothelium preserved at low temperature disappeared, then reappeared after returning to body temperature, suggesting that temperature has a direct effect on the primary cilia of corneal endothelium. [ABSTRACT FROM AUTHOR]

We report the phenotype of a 15-year-old female patient with anterior segment dysgenesis (ASD) caused by a novel heterozygous loss-of-function FOXC1 variant. The proband underwent an ophthalmic examination as well as a molecular genetic investigation comprising exome sequencing, a single nucleotide polymorphism array to access copy number and Sanger sequencing to exclude non-coding causal variants. There was bilateral mild iris hypoplasia with pupil deformation and iridocorneal adhesions. In addition to these features of ASD, the corneas were flat, with mean keratometry readings of 38.8 diopters in the right eye and 39.5 diopters in the left eye. There was a snail track lesion of the left cornea at the level of the Descemet membrane. The central corneal endothelial cell density was reduced bilaterally at 1964 and 1373 cells/mm2 in the right and left eyes, respectively. Molecular genetic analysis revealed that the proband was a carrier of a novel heterozygous frameshifting variant in FOXC1, c.605del p.(Pro202Argfs*113). Neither parent had this change, suggesting a de novo origin which was supported by paternity testing. We found no possibly pathogenic variants in the other genes associated with posterior corneal dystrophies or ASD. Further studies are warranted to verify whether there is a true association between snail track lesions, corneal flattening, and pathogenic variants in FOXC1. [ABSTRACT FROM AUTHOR]

Corneal opacities are a leading cause of visual impairment that affect 4.2 million people annually. The current treatment is corneal transplantation, which is limited by tissue donor shortages. Corneal engineering aims to develop membranes that function as scaffolds in corneal cell transplantation. Here, we describe a method for producing transplantable corneal constructs based on a collagen vitrigel (CVM) membrane and corneal endothelial cells (CECs). The CVMs were produced using increasing volumes of collagen type I: 1X (2.8 μL/mm2), 2X, and 3X. The vitrification process was performed at 40% relative humidity (RH) and 40 °C using a matryoshka-like system consisting of a shaking-oven harboring a desiccator with a saturated K2CO3 solution. The CVMs were characterized via SEM microscopy, cell adherence, FTIR, and manipulation in an ex vivo model. A pilot transplantation of the CECs/CVM construct in rabbits was also carried out. The thickness of the CVMs was 3.65–7.2 µm. The transparency was superior to a human cornea (92.6% = 1X; 94% = 2X; 89.21% = 3X). SEM microscopy showed a homogenous surface and laminar organization. The cell concentration seeded over the CVM increased threefold with no significant difference between 1X, 2X, and 3X (p = 0.323). The 2X-CVM was suitable for surgical manipulation in the ex vivo model. Constructs using the CECs/2X-CVM promoted corneal transparency restoration. [ABSTRACT FROM AUTHOR]

Myotonic dystrophy is the most common inherited muscular dystrophy in adults and presents as two forms, type 1, and type 2. Ocular manifestations such as premature cataract formation, may be the first diagnostic sign or symptom of the disease, offering ophthalmologists a unique diagnostic role. Fuchs' endothelial corneal dystrophy, ptosis and ocular melanoma are other possible findings. Systemic features can help providers better understand the disease and any accommodations to be made in clinical or surgical settings. Some patients with this disease may request evaluation of certain cataract or corneal refractive procedures. This article focuses on pertinent information for clinicians to utilize when evaluating and treating patients with myotonic dystrophy and specific surgical perspectives to consider prior to any ocular interventions. Hydrophobic intraocular lenses are still recommended in these patients with careful observation of capsular phimosis and posterior capsular opacities. [ABSTRACT FROM AUTHOR]

Corneal endothelial dystrophies are a heterogeneous group of diseases with different modes of inheritance and genetic basis for each dystrophy. The genes associated with these diseases encode transcription factors, structural components of the stroma and Descemet membrane, cell transport proteins, and others. Congenital hereditary endothelial dystrophy (CHED) is associated with mutations in two genes, OVOL2 and SLC4A11, for dominant and recessive forms of CHED, respectively. Mutations in three genes are known to cause posterior polymorphous corneal dystrophy (PPCD). They are OVOL2 (PPCD1), ZEB1 (PPCD3), and GRHL1 (PPCD4). The PPCD2 locus involving the collagen gene COL8A2 on chromosome 1 is disputed due to insufficient evidence. Mutations in the COL8A2 gene are associated with early-onset Fuchs' endothelial corneal dystrophy (FECD). Several genes have been associated with the more common, late-onset FECD. Alterations in each of these genes occur in a fraction of patients, and the most prevalent genetic alteration in FECD patients across the world is a triplet repeat expansion in the TCF4 gene. Knowledge of the genetics of corneal endothelial dystrophies has considerably advanced within the last decade and has contributed to better diagnosis of these dystrophies as well as opened up the possibility of novel therapeutic approaches based on the molecular mechanisms involved. The functions of genes identified to date provide insights into the pathogenic mechanisms involved in each disorder. [ABSTRACT FROM AUTHOR]

Introduction: Endothelial cell density (ECD) changes long after penetrating keratoplasty (PKP) of organ-cultured corneas have been little studied. We aim to calculate the point when ECD decline stabilises following PKP with organ culture stored corneas. Methods: This is an observational study of first-ever PKPs and first-ever re-grafts, performed over 17 years under a single surgeon. ECDs were acquired at 3 and 6 months, 1 year post-graft and annually thereafter by specular microscopy. Time-dependent ECD data was fitted to a log-biexponential model. Results: We studied 465 first-ever grafts and 128 re-grafts. Mean recipient age was 59 years (range 0–96 years; SD 22). Median follow-up was 5.7 (range 0.2–17.1) years. Probability of ED at 5 years in first grafts and re-grafts was 4.4% (2.6–7.1%) and 14.8% (8.3–23.2%). In first grafts, ECD loss reached 0.6% per annum at 7.9 (6.2–9.6) years post-operatively. The half-lives of ECD loss during the immediate post-operative period for first grafts, re-grafts, dystrophies, ectasias, and previous ocular surgery are 20.1 (14.9–30.9), 12.8 (6.9–79.4), 19.5 (13.1–37.7), 26.2 (16.2–68), and 11.6 (6.7–41.3) months, respectively. The half-life during this rapid phase of ECD loss has an inverse correlation with graft survival at 10 years (r = − 0.89, p = 0.02). Conclusions: Rate of endothelial decompensation is higher in first grafts than re-grafts. ECD decline stabilises 7.9 years post-operatively in first grafts but then becomes lower than the physiological loss expected. Further work is needed to verify whether organ-cultured grafts reach physiological levels of ECD loss faster than hypothermically stored grafts. [ABSTRACT FROM AUTHOR]

The corneal endothelium is the inner corneal mono-layered epithelium, fundamental for preserving corneal hydration and transparency. However, molecular mechanisms that regulate corneal endothelial cells (CEnCs), in particular regarding their proliferative capacity, have been only partially elucidated. CEnCs are quiescent in vivo and they easily undergo endothelial to mesenchymal transition (EnMT) in vitro. This study aims to analyze CEnCs behavior and expression in vitro, either in sub-confluent growing (S) or confluent (C) CEnCs cultures. Primary rabbit and human CEnCs were cultured and used for RT-PCR, immunofluorescence or western blot analysis. These methods allowed identifying a novel molecular marker, LAP2, that is upregulated in S while downregulated in C human or rabbit CEnCs. Those results were observed for several subsequent passages in culture and this, together with the correlation between ki67 and LAP2 expression, suggested LAP2 as a novel possible indicator for culture ageing. Finally, treatment with FGF and TGFβ in rCEnCs highlighted how LAP2 can vary as the cells regulate their proliferative state. In conclusion, we have identified a novel marker for CEnCs, LAP2, that regulates its expression depending on the cells sub/confluent state and that correlates with CEnCs proliferation. [ABSTRACT FROM AUTHOR]

Simple Summary: Detailed knowledge of the structure and properties of the human cornea is a prerequisite not only for the treatment of various corneal diseases but also for successful corneal transplantation and its long-term survival after grafting. Using various cell and molecular biology approaches, we found in cornea the protease inhibitor 9. This protein, known to be present in other human tissues but not yet reported in cornea, is directly involved in the immune response after transplantation. Together with its inhibitor (granzyme B), we localized this protein, especially in the superficial and inner cornea layers. This localization indicates that protease inhibitor 9 protein may be involved in protecting the cornea from external damage, but also in protection against immune cells inducing corneal graft rejection. Furthermore, we have shown on pathological corneal samples from corneal melting and herpes virus keratitis that the increased expression of both proteins is linked to these diseases. These experiments and their results represent an important contribution to the basic research of cornea biological properties with direct overlap into clinical practice. The aim of this study was to find out whether protease inhibitor 9 (PI-9) and granzyme B (GrB) molecules that contribute to immune response and the immunological privilege of various tissues are expressed in healthy and pathological human corneas. Using cryosections, cell imprints of control corneoscleral discs, we showed that PI-9 was expressed particularly in the endothelium, the superficial and suprabasal epithelium of healthy corneas, limbus, and conjunctiva. GrB was localized in healthy corneal and conjunctival epithelium, while the endothelium showed weak immunostaining. The expression of PI-6 and GrB was confirmed by qRT-PCR. Increased expression levels of the PI-9 and GrB genes were determined when the corneas were cultured with proinflammatory cytokines. Fluorescent and enzymatic immunohistochemistry of pathological corneal explants (corneal melting and herpes virus keratitis) showed pronounced PI-9, GrB, human leucocyte antigen (HLA)-DR, and leukocyte-common antigen (CD45) signals localized in multicellular stromal infiltrates and inflammatory cells scattered in the corneal stroma. We conclude that increased expression of the PI-9 and GrB proteins under pathological conditions and their upregulation in an inflammatory environment indicate their participation in immune response of the cornea during the inflammatory process. [ABSTRACT FROM AUTHOR]

Purpose: To study corneal endothelial changes post phacoemulsification in diabetic and non-diabetic patients.Methods: A comparative, prospective, observational study was conducted on 100 diabetic and 100 non-diabetics who underwent phacoemulsification. All patients were operated by the same surgeon by using the phaco chop technique to exclude any surgeon-related bias. Endothelial cell count, CCT, and coefficient of variance (CV) were measured with a specular microscope along with BCVA preoperatively and at 1 week, 4 weeks, and 3 months postoperatively. For statistical analysis, data were analyzed by using SPSS (version 27.0; SPSS Inc., Chicago, IL, USA). Data were summarized as mean and standard deviation for numerical variables and count and percentages for categorical variables. Chi square test, independent sample T test, and paired T test were used to compare the data. P ≤ 0.05 was considered statistically significant.Results: : Postoperatively at 1 week, 4-week, and 3 months follow-up intervals, the mean endothelial cell count and coefficient of variance were significantly higher, and the mean percentage of hexagonal cells was significantly lower in non diabetic as compared to the diabetic group. A significant difference in mean central corneal thickness of the two groups was observed at 1-week and 4-weeks postoperative intervals; at both these intervals, the mean value was significantly higher in non diabetic as compared to the non-diabetic group. However, at 3-months post-operative interval, the difference between the two groups was not significant statistically. Mean BCVA values were significantly higher in diabetic as compared to the diabetic group at all three follow-up intervals.Conclusion: : The findings of the present study show that endothelial cell characteristics are adversely affected in diabetic eyes as compared to non-diabetic patients undergoing phacoemulsification; this might also have an effect on the visual outcomes. [ABSTRACT FROM AUTHOR]