Hepatocyte Growth Factor Modulates Corneal Endothelial Wound Healing In Vitro.

In this study, we assessed the impact of hepatocyte growth factor (HGF) on corneal endothelial cells (CECs), finding that HGF concentrations of 100-250 ng/mL significantly increased CEC proliferation by 30%, migration by 32% and improved survival under oxidative stress by 28% compared to untreated controls (p < 0.05). The primary objective was to identify non-fibrotic pharmacological strategies to enhance corneal endothelial regeneration, addressing a critical need in conditions like Fuchs' endothelial dystrophy (FED), where donor tissue is scarce. To confirm the endothelial nature of the cultured CECs, Na+/K+-ATPase immunohistochemistry was performed. Proliferation rates were determined through BrdU incorporation assays, while cell migration was assessed via scratch assays. Cell viability was evaluated under normal and oxidative stress conditions using WST-1 assays. To ensure that HGF treatment did not trigger epithelial-mesenchymal transition, which could lead to undesirable fibrotic changes, α-SMA staining was conducted. These comprehensive methodologies provided robust data on the effects of HGF, confirming its potential as a therapeutic agent for corneal endothelial repair without inducing harmful EMT, as indicated by the absence of α-SMA expression. These findings suggest that HGF holds therapeutic promise for enhancing corneal endothelial repair, warranting further investigation in in vivo models to confirm its clinical applicability.

MitoQ relieves mitochondrial dysfunction in UVA and cigarette smoke-induced Fuchs endothelial corneal dystrophy.

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.

Neuropeptide alpha-Melanocyte stimulating hormone preserves corneal endothelial morphology in a murine model of Fuchs dystrophy.

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.

Evaluation of the effect of atorvastatin on corneal endothelial cells during the initial 12-month period after acute coronary syndrome.

PURPOSE: It was aimed to compare corneal endothelial changes during the initial 12-month period in which patients started using atorvastatin after a diagnosis of acute coronary syndrome (ACS). METHODS: Forty-six participants (group 1) who underwent cardiac angioplasty and stenting for ACS and started using 80 mg atorvastatin in the early period were included in the study. In the study, a control group comprising 71 healthy adults (group 2) was included. These individuals did not use medication for any known systemic disease, had never taken statins, had no history of ocular surgery, and did not have any cornea-related eye diseases. Baseline and 12th month endothelial evaluations of group 1 and 2 participants were compared using specular microscopy. RESULTS: There were 28 female and 18 male participants in group 1 and 48 female and 23 male participants in group 2 (P = 0.455). The mean baseline corneal endothelial cell density (CECD) was not significantly higher in group 1 compared to group 2 (2471.4 ± 200 cells/mm2 vs 2428.2 ± 539.8 cells/mm2, P = 0.230). When the change between baseline and 12th month CECD was examined, the decrease in group 2 was significantly different from that in group 1 (-15,2 ± 31,9 and -44,8 ± 49,6, P = 0,002). Although the percentage of hexagonal cells decreased significantly in group 2 participants, no significant change was observed in group 1 (respectively; P < 0.001, P = 0.073). The endothelial cell coefficient of variation did not differ significantly in group 1 participants over a 1-year period (P = 0.192), and a significant increase was observed in group 2 (P < 0.001). CONCLUSION: This study revealed that atorvastatin may have a positive effect on corneal endothelium cell density and morphology.

Protective effects of curcumin on corneal endothelial cell PANoptosis and monocyte adhesion induced by tumor necrosis factor-alpha and interferon-gamma in rats.

Decrease of human corneal endothelial cell (CEC) density leads to corneal edema, progressive corneal opacity, and reduced visual acuity. A reduction in CEC density may be related to elevated levels of inflammatory cytokines, such as tumor necrosis factor (TNF)-α and interferon (INF)-γ. PANoptosis, characterized by the activation of apoptosis, necroptosis, and pyroptosis, could be a factor in the loss of CECs driven by TNF-α and INF-γ. Cytokines also stimulate monocytes adhesion to endothelium. It has been shown in previous research that curcumin plays protective roles against numerous corneal inflammatory diseases. However, it is not determined whether curcumin acts as an anti-PANoptotic agent or if it mitigates monocyte adhesion to CECs. Therefore, this research aimed to explor the potential therapeutic effects of curcumin and its underlying mechanisms in the loss of CECs. CEC injury models were established, and curcumin was injected subconjunctivally. Clinical evaluation of the corneas was conducted using a scoring system and anterior segment photography. Corneal observation was performed with hematoxylin and eosin staining and immunostaining of zona occludens-1(ZO-1). Apoptotic cells within the corneal endothelium were observed using TUNEL staining. The detection of primary proteins expression was accomplished through Western blot analysis. Interleukin (IL)-1ß and macrophage chemotactic protein 1 (MCP-1) levels were determined via ELISA, while the expression of cleaved caspase-3, gasdermin-D (GSDMD), phosphor-mixed lineage kinase domain-like protein (p-MLKL) and intercellular cell adhesion molecule-1 were confirmed by immunofluorescence. Myeloperoxidase (MPO) activity was measured in aqueous humors. Curcumin treatment attenuated the loss of CECs and corneal edema caused by TNF-α and IFN-γ. Besides, it decreased the count of TUNEL-positive cells, and inhibited the upregulation of cleaved caspase-3, cleaved caspase-6, cleaved caspase-7, and cleaved poly (ADP-ribose) polymerase. Moreover, both the expression and phosphorylation of MLKL and receptor-interacting protein 3 were decreased in curcumin-treated rats. Furthermore, curcumin also lowered the expression of cleaved caspase-1, diminished the levels of IL1ß and MCP-1, and inhibited the activity of MPO. Besides, the expression of intercellular cell adhesion molecule-1, vascular cell adhesion molecule-1, as well as the number of CD11b-positive cells adhered to the CECs decreased for the administration of curcumin.

Laboratory exploration of the use of ripasudil in descemetorhexis with a human ex vivo model.

The aim of the study was to investigate the effect of ripasudil on corneal endothelial cell survival and migration after two types of descemetorhexis on a human ex vivo model. Eleven human corneoscleral buttons were incubated in either 50 ml organ culture medium containing 10 µM ripasudil or 50 µl dimethyl sulfoxide (DMSO), the vehicle in ripasudil for 2 days prior to wound creation then for 14 days after. The wound was created with either full trephination scoring or by shallow trephination plus manual peeling. At day 14, immunohistochemistry with vimentin and Na+/K+/ATPase markers was conducted. Tissues were assessed at day 3, 7 and 14 for morphology, cell migration, cell viability and cell density. Full trephination scoring created more damage on tissues compared to shallow trephination with full Descemet membrane peeling. In the full trephination scoring group, no differences in cell viability were noted when ripasudil and DMSO were compared. With the peeling method, Ripasudil could protect the endothelial cell death and maintain the morphology compared to the control. At day 14, no differences in the peripheral cell viability and density were found between ripasudil and DMSO, although the ripasudil group presented significantly increased central cell count and cell viability. Increased cell migration was noted with ripasudil and the initial cell morphology of those migrated cells was similar to that of fibroblasts. In conclusion, ex vivo modelling suggested that peeling resulted in less cell damage than scoring and ripasudil maintained better morphology and promoted migration. These effects might be via transformation of endothelial cells into a more motile spindle-like phenotype.

Nicotinamide promotes the differentiation of functional corneal endothelial cells from human embryonic stem cells.

Corneal transplantation represents the primary therapeutic approach for managing corneal endothelial dysfunction, but corneal donors remain scarce. Anterior chamber cell injection emerges as a highly promising alternative strategy for corneal transplantation, with pluripotent stem cells (PSC) demonstrating considerable potential as an optimal cell source. Nevertheless, only a few studies have explored the differentiation of functional corneal endothelial-like cells originating from PSC. In this investigation, a chemical-defined protocol was successfully developed for the differentiation of functional corneal endothelial-like cells derived from human embryonic stem cells (hESC). The application of nicotinamide (NAM) exhibited a remarkable capability in suppressing the fibrotic phenotype, leading to the generation of more homogeneous and well-distinctive differentiated cells. Furthermore, NAM effectively suppressed the expression of genes implicated in endothelial cell migration and extracellular matrix synthesis. Notably, NAM also facilitated the upregulation of surface marker genes specific to functional corneal endothelial cells (CEC), including CD26 (-) CD44 (-∼+-) CD105 (-) CD133 (-) CD166 (+) CD200 (-). Moreover, in vitro functional assays were performed, revealing intact barrier properties and Na+/K+-ATP pump functionality in the differentiated cells treated with NAM. Consequently, our findings provide robust evidence supporting the capacity of NAM to enhance the differentiation of functional CEC originating from hESC, offering potential seed cells for therapeutic interventions of corneal endothelial dysfunction.

Long-term dynamic changes and influencing factors of corneal morphology after multiple intravitreal injections of anti-VEGF drugs.

To observe alterations in corneal morphology caused by repeated intravitreal injections of anti-vascular endothelial growth factor (VEGF). Prospective cohort study. Seventy-seven eyes were treated with intravitreal injection of anti-VEGF from June 2021 to March 2023. There were 25 eyes of neovascular age-related macular degeneration (nAMD), 24 eyes of diabetic macular edema (DME), and 28 eyes of retinal vein occlusion (RVO). Aflibercept was used in 37 eyes and Ranibizumab was used in 40 eyes. 3 + PRN was used. Corneal endothelium and corneal thickness were measured using a corneal endothelial microscope. The data related to central corneal thickness, corneal endothelial cell density (ECD), average cell size, coefficient of variation (CV), proportion of hexagonal cells (Hex%) was collected. A comparison was also made between baseline and the dynamic changes of all indexes 1 year following the last injection. It was observed that in comparison to baseline, ECD and Hex% decreased significantly after the 3rd injection of Aflibercept and Ranibizumab. However, ECD did not decrease further and remained at the same level as after the last injection. Hex% and average cell size increased to a certain extent in comparison to the last injection. All the changes were found to be statistically significant (P < .01). After 3 injections, ECD in DME group was markedly lower than that in nAMD and RVO group, but the CV in DME group was higher than that in nAMD as well as RVO groups, and all the differences were statistically significant (P < .05). Following intravitreal anti-VEGF therapy, DME is more likely than other disorders to result in a decrease in ECD. Repeated intravitreal injections of anti-VEGF drugs can reduce the Hex% and ECD to a certain extent. After the last injection, Hex% can progressively recover, and ECD can remain stable without further declining. After injections, ECD in DME group was found to be significantly lower than that in nAMD and RVO groups, but CV in DME group was significantly higher in comparison to the other 2 groups. In patients with macular edema, repeated intravitreal injections of anti-VEGF may have certain effects on corneal morphology. Patients with diabetes mellitus in particular should pay special attention to corneal safety following repeated intravitreal injections if they have significantly reduced ECD at baseline.

Continuous Corneal Endothelial Damage by Chlorhexidine Alcohol Used for Disinfection in Nonophthalmic Surgery.

PURPOSE: To report a case of corneal endothelial damage caused by alcohol-containing chlorhexidine gluconate (CG-A) and its progression over time. METHODS: This was a case report. RESULTS: A 22-year-old man underwent neurosurgery under general anesthesia. CG-A (1%) was used for disinfection after the application of corneal protection tape. Postoperatively, the patient presented with hyperemia and swelling of the left conjunctiva and was referred to our department. Initial examination revealed left corneal epithelial erosion and corneal edema, which improved on postoperative day 14. The corneal endothelial cell density (ECD) was 3,345 cells/mm 2 on day 14, decreased rapidly to 2,090 cells/mm 2 on day 42, and slowly reduced to 1,122 cells/mm 2 on day 168. Thereafter, no decrease in ECD was observed. CONCLUSIONS: CG formulations can lead to a persistent decrease in ECD over several months, even after improvement of acute corneal edema.

"Keep on ROCKIn": Repurposed ROCK inhibitors to boost corneal endothelial regeneration.

The global shortage of corneal endothelial graft tissue necessitates the exploration of alternative therapeutic strategies. Rho-associated protein kinase inhibitors (ROCKi), recognized for their regenerative potential in cardiology, oncology, and neurology, have shown promise in corneal endothelial regeneration. This study investigates the repurposing potential of additional ROCKi compounds. Through screening a self-assembled library of ROCKi on B4G12 corneal endothelial cells, we evaluated their dose-dependent effects on proliferation, migration, and toxicity using live-cell imaging. Nine ROCKi candidates significantly enhanced B4G12 proliferation compared to the basal growth rate. These candidates were further assessed for their potential to accelerate wound closure as another indicator for tissue regeneration capacity, with most demonstrating notable efficacy. To assess the potential impact of candidate ROCKi on key corneal endothelial cell markers related to cell proliferation, leaky tight junctions and ion efflux capacity, we analyzed the protein expression of cyclin E1, CDK2, p16, ZO-1 and Na+/K+-ATPase, respectively. Immunocytochemistry and western blot analysis confirmed the preservation of corneal endothelial markers post-treatment with ROCKi hits. However, notable cytoplasm enlargement and nuclear fragmentation were detected after the treatment with SR-3677 and Thiazovivin, indicating possible cellular stress. In compared parameters, Chroman-1 at a concentration of 10 nM outperformed other ROCKi, requiring significantly 1000-fold lower effective concentration than established ROCKi Y-27632 and Fasudil. Altogether, this study underscores the potential of repurposing ROCKi for treating corneal endothelial dysfunctions, offering a viable alternative to conventional grafting methods, and highlights Chroman-1 as a promising candidate structure for hit-to-lead development.

The effects of trypan blue use on the corneal endothelium during cataract surgery in patients with pseudoexfoliation syndrome (PEX).

PURPOSE: In the present clinical study, it was aimed to investigate the possible effects of Trypan blue (TB) use on the corneal endothelium during cataract surgery in eyes with pseudoexfoliation syndrome (PEX) during a three-month follow-up period using the contralateral eye control design. METHODS: This prospective, randomised controlled, individual cohort study included 92 eyes of 46 patients with bilateral PEX and cataracts. While 1% TB was applied to one eye of the patients before capsulorhexis (study group), it was not applied to the other eye (control group). Both groups were compared preoperatively and postoperatively in terms of endothelial cell density (ECD), endothelial cell loss (%), pleomorphism, polymegathism and central corneal thickness (CCT) using specular microscopy. RESULTS: Preoperative corneal ECD was measured as 2362.56 ± 253.27 in the study group, 2380.84 ± 220.54 in the control group, and 2145.58 ± 221.71 in the study group and 2184.97 ± 200.94 cells/mm2 in the control group in the postoperative 3rd-month follow-up (p = 0.71 and = 0.37, respectively). In addition, there were no significant differences between the two groups in terms of the percentage of hexagonal cells, coefficient of variation (CV), and CCT both preoperatively and postoperatively 3 months later (p = 0.78, =0.39, =0.95 preoperatively and p = 0.31, =0.26, =0.83 postoperatively, respectively). CONCLUSION: This study demonstrated that the injection of 1% TB into the anterior chamber for staining the anterior capsule during cataract surgery did not cause significant corneal endothelial changes at postoperative 3rd months, despite the increased fragility of corneal endothelial cells in patients with PEX.

Effect of Rho-Associated Kinase Inhibitor and Mesenchymal Stem Cell-Derived Conditioned Medium on Corneal Endothelial Cell Senescence and Proliferation.

This study aims to obtain sufficient corneal endothelial cells for regenerative application. We examined the combinatory effects of Rho-associated kinase (ROCK) inhibitor Y-27632 and mesenchymal stem cell-derived conditioned medium (MSC-CM) on the proliferation and senescence of rabbit corneal endothelial cells (rCECs). rCECs were cultured in a control medium, a control medium mixed with either Y-27632 or MSC-CM, and a combinatory medium containing Y-27632 and MSC-CM. Cells were analyzed for morphology, cell size, nuclei/cytoplasmic ratio, proliferation capacity and gene expression. rCECs cultured in a combinatory culture medium showed a higher passage number, cell proliferation, and low senescence. rCECs on collagen type I film showed high expression of tight junction. The cell proliferation marker Ki-67 was positively stained either in Y-27632 or MSC-CM-containing media. Genes related to cell proliferation resulted in negligible changes in MKI67, CIP2A, and PCNA in the combinatory medium, suggesting proliferative capacity was maintained. In contrast, all of these genes were significantly downregulated in the other groups. Senescence marker ß-galactosidase-positive cells significantly decreased in either MSC-CM and/or Y-27632 mixed media. Senescence-related genes downregulated LMNB1 and MAP2K6, and upregulated MMP2. Cell cycle checkpoint genes such as CDC25C, CDCA2, and CIP2A did not vary in the combinatory medium but were significantly downregulated in either ROCK inhibitor or MSC-CM alone. These results imply the synergistic effect of combinatory culture medium on corneal endothelial cell proliferation and high cell number. This study supports high potential for translation to the development of human corneal endothelial tissue regeneration.

Corneal endothelial cell density in patients receiving chemotherapy.

PURPOSE: This study aimed to determine if the corneal endothelium was affected by chemotherapy. METHODS: Chemotherapy patients were recruited to undergo specular microscopy before treatment and again at 1- and 2-year follow-up visits. One eye per patient, per follow-up, was selected for comparison to baseline. RESULTS: Forty-six volunteers completed baseline and at least one follow-up assessment. From 51 eyes, there was no significant change in endothelial cell density for 41 eyes assessed at one year (MD = 0.73%, 95% CI -1.33 to 2.78%) and 18 eyes at two years (MD = 0.31%, 95% CI -3.53 to 4.15%). CONCLUSION: Although other studies have shown that chemotherapy can adversely affect the corneal epithelium, this study showed no measurable change in endothelial cell density.

Tumor necrosis factor-alpha and interferon-gamma induce inflammasome-mediated corneal endothelial cell death.

PURPOSE: Chronic corneal endothelial cell (CEC) loss results in corneal edema and vision loss in conditions such as pseudophakic bullous keratopathy (PBK), Fuchs' dystrophy, and corneal graft failure. Low CEC density has been associated with an elevation of intraocular pro-inflammatory cytokines such as tumor necrosis factor (TNF)-α and interferon (INF)-γ. These cytokines are capable of triggering pyroptosis, a programmed cell death mechanism mediated by the inflammasome, prompting the activation of the pro-inflammatory cytokine interleukin (IL)-1ß, the perpetuation of inflammation, and subsequent damage of corneal endothelial tissue. Therefore, the purpose of this study was to determine the deleterious contribution of the inflammasome and pyroptosis to CEC loss. METHODS: CECs from human donor corneas were treated ex vivo with TNF-α and IFN-γ for 48 h. Levels of caspase-1 and IL-1ß were then assayed by ELISA, and the expression of caspase-1 and gasdermin-D (GSDM-D) were confirmed by immunofluorescence. Endothelial cell damage was analyzed by a lactate dehydrogenase (LDH) release assay, and oxidative stress was determined by measuring the levels of reactive oxygen species (ROS) in the culture media. RESULTS: Inflammasome activation and oxidative stress were elevated in CECs following exposure to TNF-α and IFN-γ, which resulted in cell death by pyroptosis as determined by LDH release which was inhibited by the caspase-1 inhibitor Ac-YVAD-cmk. CONCLUSION: CEC death is induced by the pro-inflammatory cytokines TNF-α and IFN-γ, which contribute to inflammasome activation. Moreover, the inflammasome is a promising therapeutic target for the treatment of chronic CEC loss.

The effect of intravitreal anti-vascular endothelial growth factor injections on corneal endothelium in patients with diabetic macular oedema.

PURPOSE: To evaluate the characteristics of corneal parameters in patients with diabetic macular oedema (DME) treated with intravitreal anti-vascular endothelial growth factor (anti-VEGF) injections. METHODS: Participants in this study were 36 patients with DME, treated with either intravitreal ranibizumab (n = 16) or aflibercept (n = 20). All participants underwent best-corrected visual acuity (BCVA) measurement, optical coherence tomography and non-contact specular microscopy to evaluate corneal endothelium parameters (endothelial cell density-ECD, hexagonality, coefficient of variation of the cell size and central corneal thickness-CCT), at baseline and at months 6 and 12 after the first intravitreal injection. Comparisons between baseline and months 6 and 12 were performed. RESULTS: There was no statistically significant difference regarding ECD, hexagonality, coefficient of variation of the cell size and CCT at month 6 and 12 post initial injection compared to baseline in patients with DME. BCVA improved significantly at month 6 and 12 compared to baseline (p < 0.001 for both comparisons). Central retinal thickness was significantly reduced at month 6 and 12 compared to baseline (p < 0.001 for both comparisons). CONCLUSION: Intravitreal anti-VEGF injections in patients with DME were found not to affect corneal parameters, namely ECD, hexagonality, coefficient of variation of the cell size and CCT at the long-term follow-up of 12 months.

Cytomegalovirus endotheliitis with recurrent anterior uveitis and secondary glaucoma misdiagnosed as HLA-B27 uveitis.

A 61-year-old male patient presented with decreased vision and recurrent redness in his right eye since the past 4 years. He had been diagnosed elsewhere as HLA-B27 positive anterior uveitis and was on oral methotrexate and topical corticosteroids for recurrent disease. He was on maximal medical therapy for glaucoma. Examination showed prominent inferior corneal oedema with pigmented keratic precipitates and elevated intraocular pressure. He underwent combined trabeculectomy with mitomycin C and cataract surgery. The aqueous sample tested positive for cytomegalovirus. He responded well to oral valganciclovir with resolution of uveitis, the intraocular pressure was well controlled and the corneal oedema resolved completely.

Multiple roles of FGF10 in the regulation of corneal endothelial wound healing.

Corneal endothelial dysfunction usually induces corneal haze and oedema, which seriously affect visual function. The main therapeutic strategy for this condition is corneal transplantation, but the use of this strategy is limited by the shortage of healthy donor corneas. Compared with corneal transplantation, drug intervention is less invasive and more accessible; thus, finding an effective pharmaceutical alternative for cornea transplantation is critical for the treatment of corneal endothelial dysfunction. In this study, we established a rabbit scratch model to investigate the effect of fibroblast growth factor 10 (FGF10) on corneal endothelial wound healing. Results showed that FGF10 injection accelerated the recovery of corneal transparency and increased the protein expression levels of ZO1, Na+/K+-ATPase and AQP-1. Moreover, FGF10 significantly inhibited the expression levels of endothelial-to-mesenchymal transition proteins and reduced the expression levels of the proinflammatory factors IL-1ß and TNF-α in the anterior chamber aqueous humour. FGF10 also enhanced the Na+/K+-ATPase activity by enhancing mitochondrial function as a result of its direct interaction with its conjugate receptor. Thus, FGF10 could be a new pharmaceutical preparation as treatment for corneal endothelial dysfunction.

Optimizing Visualization of Descemet Membrane Endothelial Keratoplasty Tissue: Assessing the Impact of Trypan Blue Exposure on Stain Duration and Corneal Endothelial Cell Function.

PURPOSE: To assess how trypan blue staining affects Descemet membrane endothelial keratoplasty (DMEK) graft visibility and corneal endothelial cell (CEC) mitochondrial respiration. METHODS: DMEK grafts (n = 20) were stained with trypan blue 0.06% for 1, 3, 5, or 10 minutes. Each graft was injected into an artificial anterior chamber. Surgery was simulated with tapping and sweeping motions on the corneal surface and injections of balanced salt solution (BSS). Graft visibility was assessed at 5, 10, 20, and 30 minutes. Effects of trypan blue on mitochondrial respiration were assessed using primary CECs cultured from donor corneas (n = 43). Treatment wells exposed to trypan blue 0.06% (1, 5, or 30 minutes) and donor-matched control wells to methylene blue 1% (1 minute) or BSS (1, 5, or 30 minutes) were assayed for key respiration parameters. RESULTS: After 5 minutes of surgical manipulation, grafts stained for 5 minutes were significantly more visible than grafts stained for 1 or 3 minutes; there was no added benefit of staining for 10 minutes. After 10 minutes of surgical manipulation, grafts stained for 3 minutes were more visible than grafts stained for 1 minute, without additional benefits of staining ≥5 minutes. No visibility differences were observed after ≥20 minutes of surgical manipulation. CEC mitochondrial respiration did not change significantly following trypan blue exposure for all intervals tested compared to BSS. CONCLUSIONS: Staining DMEK grafts with trypan blue for 3 to 5 minutes optimizes visibility during surgical manipulation without mitochondrial impairment. Corneal surgeons learning DMEK will benefit from optimizing this critical step.

Endothelial Biopsy for the Diagnosis and Management of Culture-Negative Retrocorneal Fungal Keratitis With the Assistance of Optical Coherence Tomography Imaging.

PURPOSE: To report a case of a culture-negative deep fungal corneal infection that was diagnosed after histopathology of an anterior segment optical coherence tomography-guided endothelial biopsy. METHODS: A 22-year-old woman with history of contact lens wear and concomitant topical steroid use presented with a mid-stromal corneal infiltrate that failed to respond to oral acyclovir and topical fortified antibiotics. Although cornea stains, cultures, and confocal microscopy showed negative results, there was high clinical suspicion for fungal keratitis. After 2 months on topical natamycin, oral voriconazole, and serial intrastromal and intracameral voriconazole injections, the infiltrate enlarged and deepened. Imaging with anterior segment optical coherence tomography revealed that the infection had progressed to an endothelial plaque. RESULTS: Diagnostic endothelial biopsy was performed in the operating room. Cultures showed again negative results, whereas histopathology of the removed specimen revealed fungal elements. The postoperative edema at the site of the biopsy resolved over the course of 4 weeks, and a posterior stromal scar formed. Serial intrastromal and intracameral voriconazole injections were continued for the first postoperative month. At the 1-year and the 3-year follow-up examinations, the patient's vision was 20/20 without recurrence. CONCLUSIONS: Intraoperative scraping of the endothelial plaque and histopathologic evaluation of the specimen proved to be of utmost importance for definitive diagnosis and resolution of the culture-negative deep fungal infection in this case. This young patient's cornea was retained and vision remains excellent.

Effect of Intracameral Cefuroxime on Graft Endothelial Cells in Descemet Membrane Endothelial Keratoplasty: A Comparative Study.

Purpose: To evaluate the effect of intracameral cefuroxime on graft endothelial cell loss after simple Descemet Membrane Endothelial Keratoplasty (DMEK) and combined DMEK and cataract surgery.Materials and Methods: Single-center retrospective comparative analysis. One hundred and three patients were included, 31 in the cefuroxime group and 72 in the non-cefuroxime (NC) group. Best Spectacle-Corrected Visual Acuity (BSCVA), endothelial cell density (ECD) of the graft measured by specular microscopy, and the recipient's pachymetry were recorded pre-operatively and at 1, 3, and 6 months after surgery.Results: In the cefuroxime group, BSCVA was 0.22 ± 0.27 LogMAR, 0.15 ± 0.24 LogMAR and 0.07 ± 0.22, respectively, at 1, 3, and 6 months after surgery with no significant differences found when compared to the NC group (p > .05). Anatomical outcomes were similar as mean pachymetry decreased from 599 ± 51 µm preoperatively to 511 ± 30 µm at 6 months after surgery in the cefuroxime group and from 607 ± 67 µm preoperatively to 519 ± 32 µm at 6 months in the NC group (p = .25). Endothelial cell loss was comparable between both groups: 33.4% versus 33.6% at 1 month (p = .97), 37.4% versus 34.9% at 3 months (p = .68) and 41.6% versus 38.3% at 6 months (p = .42) in the cefuroxime and NC groups, respectively. The rates of rebubbling, graft rejection, and cystoid macular edema were not significantly higher in the cefuroxime group.Conclusion: The use of intracameral cefuroxime during simple or combined DMEK did not lead to higher graft endothelial cell loss.