This systematic review and meta-analysis evaluated available data on the diagnostic accuracy of ex vivo confocal laser scanning microscopy (EVCM) for the diagnosis of basal cell carcinoma (BCC) in tissue specimens and for margin assessment in micrographic surgery, using conventional histopathological analysis as reference standard. MEDLINE and Embase were systematically searched in November 2024 in accordance with PRISMA guidelines to identify diagnostic accuracy studies evaluating EVCM for BCC. Pooled sensitivity and specificity were estimated using bivariate random effects models. Sixteen studies comprising 1,387 specimens were included. The pooled mean EVCM acquisition time was 7.0 min (95% confidence interval [CI]: 5.3-9.4). For BCC diagnosis in tissue specimens, pooled sensitivity was 91.4% (95% CI: 85.9-94.9) and pooled specificity was 91.1% (95% CI: 79.2-94.5). Regarding margin assessment, pooled sensitivity and specificity were 79.5% (95% CI: 65.0-89.0) and 92.2% (95% CI: 85.5-95.9). In conclusion, EVCM shows high sensitivity and specificity for BCC diagnosis in tissue specimens and may selectively replace conventional histology when rapid diagnosis is desired. For margin assessment, EVCM has high specificity but moderate sensitivity. Therefore, conventional histology should still be considered as a subsequent step, particularly after the final excision stage of stepwise micrographic surgery.

Background and Objectives: The objective of this study is to evaluate whether deferoxamine modulates cell biological properties, such as proliferation and wound closure of porcine corneal endothelial cells (CECs) in vitro, and whether the treatment of CECs with deferoxamine results in an enhanced expression of vascular endothelial growth factor (VEGF). Materials and Methods: Corneal endothelial cells were extracted from porcine globes within 24 h postmortem. Immunohistochemistry for the endothelial Na+/K+-ATPase was performed to confirm the cells' endothelial origin. To assess CEC viability and proliferation, a water-soluble tetrazolium salt (WST-1) and 5-bromo-2'-deoxyuridine (BrdU) assay were performed. Corneal endothelial wound closure was evaluated using a wound closure assay. VEGF mRNA expression was evaluated using real-time polymerase chain reaction (rt-PCR). Results: The extracted corneal endothelial cells showed a typical hexagonal morphology with Na+/K+-ATPase staining of the cell membrane. The treatment with 200 µM deferoxamine significantly increased CEC viability to 121 ± 24% compared to the control group (p = 0.0024). Corneal endothelial cell proliferation did not show any significant changes under the treatment with deferoxamine (p > 0.05). Both 100 µM and 200 µM deferoxamine led to a significantly smaller remaining wound area of 82.4 ± 6.7% and 78.7 ± 6.2% (p < 0.0001) in comparison to the control group after 24 h of treatment in the wound closure assay. Treatment with 200 µM deferoxamine significantly induced VEGF mRNA expression to 1.67- ± 0.57-fold from 1.00- ± 0.03-fold in the control group (p = 0.0006). Conclusions: Deferoxamine effectively enhances corneal endothelial cell viability and wound healing associated with an overexpression of VEGF. Thus, deferoxamine is a potent modulator of cell biological properties of corneal endothelial cells and maintains their integrity in vitro.

The regular inspection of the eye health is an essential part of the veterinary care task in an animal facility in order to achieve robust results during experimentation. The aim of the study was to find out, when and how ocular diseases are recorded in laboratory animal facilities and what measures are taken. An online questionnaire was sent to people who work in a research environment and are involved in monitoring of animal wellbeing. In addition, an evaluation of 142 veterinary clinical cases in mice were carried out under routine conditions with the help of ophthalmological tools. 128 participants from three European countries took part in the survey. 93% stated, that eye changes, if recognized, are recorded during cage changing but in 70% of the responses, there was no systematic analysis of the underlying causes. The population study showed that technical aids are indispensable for the detection of eye lesions in mice. The survey and the population study show that although recommended, routine checks of the eye are rarely carried out and usually without suitable technical aids. From the point of view of good veterinary practice, more attention should be paid to the ocular health in laboratory animal husbandry.

OBJECTIVE:Analyzing characteristics of ocular surface squamous cell neoplasia (OSSN) at first diagnosis and potential risk factors for aggressive growth behaviour. DESIGN:Retrospective. METHODS:Including patients with first diagnosis of OSSN at a tertiary center from 2013 until 2022. Cases were analyzed regarding demographics, clinical findings, and histopathological findings, including Ki-67 expression. RESULTS:A total of 153 patients with first diagnosis of histopathological confirmed OSSN were included. Mean age was 72 years (36-98), with a slight male predominance (66%; n = 101). Most patients had invasive squamous cell carcinoma (SCC; 45.8%, 70), followed by carcinoma in situ (CIS; 37.9%, 58) and epithelial dysplasia (ED; 16.3%, 25). Duration of symptoms varied significantly: ED 6 months (0-36), CIS 1.5 (0-48), SCC 3 (0-36) (p = 0.048). 44.3% (51/115) of cases were previously misdiagnosed, and, therefore, inadequately treated. Orbital involvement was observed in 8.5% (13), intraocular in 1.3% (2), metastasis in 2.7% (4) at initial diagnosis. Ki-67 labeling index (LI) varied significantly across subtypes: ED 35% (2-87%), CIS 45% (11-85%), SCC 50% (18-93%) (p = 0.007) and was higher with involvement of the caruncle, lower fornix, lower eyelid margin, or tarsus (p = 0.023). Patients with globe or orbit invasion had significantly longer median symptom duration (6 months (0-48) vs 2 (0-48); p = 0.01). Patients with metastasis exhibited significantly higher Ki-67 LI (p = 0.027). CONCLUSIONS:Our study found extended time intervals from first symptoms to first correct diagnosis correlate with higher risk for advanced SCC. Further, elevated Ki-67 LI correlated with more invasive tumor entities, such as SCC and CIS, and indicate an increased risk of metastasis.

Objective: Ex vivo confocal laser scanning microscopy (EVCM) is an emerging imaging technique, which offers rapid tissue examination. While the current literature shows promising results in the evaluation of non-melanoma skin cancer, only limited research exists on the application of EVCM in melanocytic lesions. This study aimed to assess the utility of EVCM in the characterization of melanocytic lesions and compare its findings with gold-standard histopathology. Methods: A total of 130 skin lesions, including 76 benign and 54 malignant melanocytic lesions, were prospectively collected and imaged using EVCM. Three blinded investigators were asked to identify characteristic morphologic features observed in the lesions and classify them into benign vs. malignant. The results were then compared with the corresponding histopathology. Sensitivity and specificity were calculated using contingency tables to assess the diagnostic performance. Results: The application of EVCM allowed for the visualization of cellular and tissue-level details, including cellular pleomorphism and atypical melanocytes. A comprehensive list of benign and malignant features identified by EVCM was compiled. Using these diagnostic criteria, the imaging of the inexperienced and dermatohistopathology-experienced investigator reached 67.7% concordance, and the imaging trained dermatologist obtained 69.2% agreement with dermatohistopathology in differentiating benign vs. malignant lesions. The imaging-trained dermatohistopathologist performed best with concordance up to 79.2%. Conclusions: In conclusion, EVCM is a promising technique for the rapid assessment of melanocytic lesions. Our study provides a comprehensive overview of morphologic EVCM features, which will contribute to the development of diagnostic algorithms for accurate diagnosis and appropriate treatment planning. Further studies are needed to evaluate its clinical utility and validate our diagnostic criteria.

Galectin-3 is a multifunctional protein that is associated with diseases of the chorioretinal interface, in which the retinal pigment epithelium (RPE) plays a central role in disease development and progression. Since galectin-3 can function extracellularly as well as intracellularly via different mechanisms, we developed an immortalized human RPE cell line (ARPE-19) with a knockdown for galectin-3 expression (ARPE-19/LGALS3+/−) using a sgRNA/Cas9 all-in-one expression vector. By Western blot analysis, a reduced galectin-3 expression of approximately 48 to 60% in heterozygous ARPE-19/LGALS3+/− cells was observed when compared to native controls. Furthermore, ARPE-19/LGALS3+/− cells displayed a flattened, elongated phenotype with decreased E-cadherin as well as enhanced N-cadherin and α-smooth muscle actin mRNA expression, indicating an epithelial–mesenchymal transition of the cells. Compared to wildtype controls, ARPE-19/LGALS3+/− cells had significantly reduced metabolic activity to 86% and a substantially decreased proliferation to 73%. Furthermore, an enhanced cell adhesion and a diminished migration of immortalized galectin-3 knockdown RPE cells was observed compared to native ARPE-19 cells. Finally, by Western blot analysis, reduced pAKT, pERK1/2, and β-catenin signaling were detected in ARPE-19/LGALS3+/− cells when compared to wildtype controls. In summary, in RPE cells, endogenous galectin-3 appears to be essential for maintaining the epithelial phenotype as well as cell biological functions such as metabolism, proliferation, or migration, effects that might be mediated via a decreased activity of the AKT, ERK1/2, and β-catenin signaling pathways.

Galectins have the potential to interact with transmembrane glycoproteins to modulate their functions. Since galectin-1 interacts with PDGF-Rβ, we analyzed the effect of galectin-1 on PDGF-BB-mediated AKT signaling in primary human retinal pigment epithelial (RPE) cells and galectin-1-deficient immortalized human RPE cells (LGALS1−/−/ARPE-19) following incubation with PDGF-BB and galectin-1. Expression and localization of galectin-1, PDGF-Rβ and pAKT were investigated using western blot analysis and immunohistochemical staining. Cell proliferation of RPE cells was analyzed using BrdU ELISA. Following treatment of human RPE cells with human recombinant (hr)-galectin-1 and PDGF-BB, an intense clustering of PDGF-Rβ and colocalization with galectin-1 were detected. By Western blot analysis and immunocytochemistry of human RPE cells, an enhanced PDGF-BB-mediated expression of pAKT was observed, which was substantially reduced by additional incubation with hr-galectin-1. Vice versa, in LGALS1−/−/ARPE-19 cells, the PDGF-BB-induced pAKT signal was enhanced compared to wild-type cells. Furthermore, a decreased expression of PDGF-Rβ in human RPE cells was observed after treatment with PDGF-BB and hr-galectin-1, while in untreated LGALS1−/−/ARPE-19 cells, its constitutive expression was increased. In addition, after treatment of RPE cells with hr-galectin-1, the PDGF-BB-induced proliferation was markedly reduced. In summary, galectin-1 has the distinct potential to reduce PDGF-mediated pAKT signaling and proliferation in human RPE cells—an effect that is most likely facilitated via a decreased expression of PDGF-Rβ.

Fehlbildungen und Dystrophien der Uvea sind seltene Erkrankungen des Auges, denen Störungen in der Augenentwicklung vorausgegangen sind und die durch spezifische genetische Defekte verursacht werden können. Um deren Pathogenese besser zu verstehen, soll im ersten Abschnitt die physiologische Entwicklung der Uvea und der angrenzenden Gewebe kurz rekapituliert werden. Nachfolgend wird auf die Pathogenese, das Krankheitsbild sowie auf mögliche Therapieansätze der klinisch relevantesten Fehlbildungen und Dystrophien der Uvea spezifisch eingegangen.

Introduction The intraocular lens (IOL) can be used as a slow-release drug carrier in cataract surgery to alleviate posterior capsular opacification (PCO). The following is a systematic development of an IOL using methotrexate and the solvent casting process with poly (lactic-co-glycolic acid) (PLGA) as a carrier polymer. Methods Different solvents for PLGA and methotrexate were tested for dissolution properties and possible damage to the IOL. The required biological concentration of methotrexate was determined in human capsular bags implanted with an IOL. To detect fibrosis, α-SMA, f-actin, and fibronectin were labelled by immunofluorescence staining. Cell proliferation and extracellular matrix contraction were observed in a lens epithelial cell line (FHL-124). Finally, the IOL was designed, and an ocular pharmacokinetic model was used to measure drug release. Results Solvent mixtures were found to allow coating of the IOL with drug and PLGA without damaging it. PCO in the capsular bag model was inhibited above 1 μM methotrexate ( p = 0.02). Proliferation in FHL-124 was significantly reduced above a concentration of 10 nM ( p = 0.04) and matrix contraction at 100 nM ( p = 0.02). The release profile showed a steady state within therapeutic range. Conclusion After determination of the required physicochemical manufacturing conditions, a drug releasing IOL was designed. A favourable release profile in an ocular pharmacokinetics model could be shown.

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.

AIM: To investigate a novel phacoemulsification system “EVA NEXUS” (D.O.R.C., Dutch Opthalmic Research Center) in comparison to the existing system “EVA” in clinical use. And to compare both phacoemulsification systems in terms of efficiency, safety and postoperative inflammatory activity. METHODS: In this study standardized cataract surgery was performed on both eyes of the study participant, using the “EVA system” (control group, n=20) on one eye and the “EVA NEXUS system” (intervention group, n=20) on the other eye. Only patients with cataract LOCS Grading 1-3 and no accompanying eye diseases were included in this study. A total of 20 patients were included in this study, with each treatment arm including 20 eyes. During surgery a 0.1 mL aqueous humor sample was collected 1min after phacoemulsification to measure the total prostaglanin E2 concentrations using an enzyme-linked immunosorbent assay. The endothelial cell count, visual and refractive outcomes, and anterior chamber flare were evaluated preoperatively, and 1d, 1wk, and 3mo postoperatively. RESULTS: There were no statistically significant differences between both groups regarding intraoperative safety parameters including effective phacoemulsification time (P=0.904), balanced saline solution flow (P=0.701) and total surgery time (P=0.565). Postoperative prostaglandin E2 levels, anterior chamber flare as well as endothelial cell loss tended to be lower in the NEXUS-Group, however not being statistically significant (P=0.718; 0.164; 0.486). Both systems provided similar clinical outcomes, regarding best corrected visual acuity and refractive parameters, showing no statistically significant differences between both groups. CONCLUSION: Both systems show a high level of safety and efficency with similar results in terms of safety parameters including postoperative inflammatory activity and endothelial cell loss as well as visual and refractive outcomes. Although statistically not significant, the EVA NEXUS system tends to cause less postoperative inflammation with lower prostaglandin E2 levels and lower anterior chamber flare values.

In the eye, an increase in galectin-1 is associated with various chorioretinal diseases, in which retinal pigment epithelium (RPE) cells play a crucial role in disease development and progression. Since little is known about the function of endogenous galectin-1 in these cells, we developed a galectin-1-deficient immortalized RPE cell line (ARPE-19-LGALS1−/−) using a sgRNA/Cas9 all-in-one expression vector and investigated its cell biological properties. Galectin-1 deficiency was confirmed by Western blot analysis and immunocytochemistry. Cell viability and proliferation were significantly decreased in ARPE-19-LGALS1−/− cells when compared to wild-type controls. Further on, an increased attachment of galectin-1-deficient RPE cells was observed by cell adhesion assay when compared to control cells. The diminished viability and proliferation, as well as the enhanced adhesion of galectin-1-deficient ARPE-19 cells, could be blocked, at least in part, by the additional treatment with human recombinant galectin-1. In addition, a significantly reduced migration was detected in ARPE-19-LGALS1−/− cells. In comparison to control cells, galectin-1-deficient RPE cells had enhanced expression of sm-α-actin and N-cadherin, whereas expression of E-cadherin showed no significant alteration. Finally, a compensatory expression of galectin-8 mRNA was observed in ARPE-19-LGALS1−/− cells. In conclusion, in RPE cells, endogenous galectin-1 has crucial functions for various cell biological processes, including viability, proliferation, migration, adherence, and retaining the epithelial phenotype.

Proliferative vitreoretinopathy (PVR) as a rare fibrotic ocular disease is the main reason for failure of retinal detachment surgery and a reduced prognosis following surgery. Amniotic membrane (AM) is a versatile surgical tool for tissue stabilization, antifibrotic properties, and regeneration. Initial clinical case studies now demonstrated intravitreal tolerance as well as good anatomical and functional results for degenerative retinal diseases. Due to its diverse wound healing properties, AM could have promoting, suppressive, or no effects on PVR. To illuminate the potential of epiretinal AM transplantation in complex retinal detachment cases, we investigated its influence on human primary PVR (hPVR) cells in vitro. In our cell culture study, hPVR cells were isolated from surgically removed PVR membranes. Following incubation with AM for 48 h, AM-incubated hPVR showed significantly reduced proliferation (BrdU-ELISA; p < 0.001), migration (Boyden chamber, scratch assay; p = 0.003 and p < 0.001), and cell adhesion (p = 0.005). Collagen contraction was nearly unaffected (p = 0.04), and toxicity (histone-complexed DNA ELISA, WST-1 assay, and life/dead staining) was excluded. Next, immunofluorescence showed a myofibroblastic phenotype with reduced expression of fibrosis markers in AM-incubated cells, which was confirmed by Western blot analysis. In the proteomics assay, AM significantly regulated proteins by a more than 2-fold increase in expression which were related to the cytoskeleton, lipid metabolism, cell-matrix contraction, and protein folding. In conclusion, this in vitro work suggests no induction of fibrosis and other key steps in the pathogenesis of PVR through AM but rather inhibiting properties of profibrotic cell behavior, making it a possible candidate for suppression of PVR. Further clinical studies are necessary to evaluate the therapeutic relevance.

Amniotic membrane (AM) is a popular treatment for external ocular diseases. First intraocular implantations in other diseases reported promising results. Here, we review three cases of intravitreal epiretinal human AM (iehAM) transplantation as an adjunct treatment for complicated retinal detachment and analyze clinical safety. Possible cellular rejection reactions against the explanted iehAM were evaluated and its influence was assessed on three retinal cell lines in vitro. Three patients with complicated retinal detachment and implanted iehAM during pars plana vitrectomy are retrospectively presented. After removal of the iehAM at subsequent surgery, tissue-specific cellular responses were studied by light microscopy and immunohistochemical staining. We investigated the influence of AM in vitro on retinal pigment epithelial cells (ARPE-19), Müller cells (Mio-M1), and differentiated retinal neuroblasts (661W) . An anti-histone DNA ELISA for cell apoptosis, a BrdU ELISA for cell proliferation, a WST-1 assay for cell viability, and a live/dead assay for cell death were performed. Despite the severity of the retinal detachment, stable clinical outcomes were obtained in all three cases. Immunostaining of the explanted iehAM showed no evidence of cellular immunological rejection. In vitro, there was no statistical significant change in cell death or cell viability nor were proliferative effects detected on ARPE-19, Müller cells, and retinal neuroblasts exposed to AM. iehAM was a viable adjuvant with many potential benefits for treatment of complicated retinal detachment. Our investigations could not detect any signs of rejection reactions or toxicity. Further studies are needed to evaluate this potential in more detail.

Vaskuläre Läsionen der Bindehaut sind eine Gruppe seltener Erkrankungen, bei denen es zu einem abnormen Wachstum oder einer abnormen Entwicklung von Blutgefäßen oder Lymphgefäßen in der Bindehaut kommt (1-5). Die Gefäße können erweitert, gewunde oder missgebildet sein, die Rötungen, Schwellungen und Blutungen im betroffenen Bereich verursachen können (1-5). Sie können grob in zwei Hauptkategorien eingeteilt werden: vaskuläre Tumore wie Hämangiome und vaskuläre Fehlbildungen wie Hamartome oder lymphatisch-venöse Fehlbildungen (LVM), früher als Lymphangiome bekannt (1-6). Vaskuläre Tumoren zeichnen sich durch schnell wachsende, gut abgegrenzte Massen von Blutgefäßen aus, während vaskuläre Fehlbildungen in der Regel schon bei der Geburt vorhanden sind und aus abnormal geformten Blutgefäßen bestehen, die nicht wie Tumoren wuchern (6). Neben diesen gutartigen Tumoren und Fehlbildungen ist das Kaposis-Sarkom ein seltener bösartiger Tumor (2). Alle diese Läsionen können ein oder beide Augen betreffen und Symptome wie verschwommenes Sehen, Fremdkörpergefühl, Rötung, Epiphora und Blutungen verursachen. Die Behandlungsstrategien hängen von der Ätiologie, der Größe und der Lage der Läsion sowie den Symptomen des Patienten ab und können unter anderem Beobachtung, chirurgische Entfernung, Laser- und Kryotherapie sowie Injektion von Therapeutika umfassen. (1-9). Wir beschreiben einen Patienten mit einer isolierten konjunktivalen LVM, der sich mit zystischem Hyposphagma in unserer Notaufnahme vorstellte. Vascular lesions of the conjunctiva are a group of rare conditions that involve abnormal growth or development of blood vessels or lymphatics in the conjunctiva (1-5). They may present as dilated, tortuous, or malformed vessels, which can cause redness, swelling, and bleeding in the affected area (1-5). They can be broadly classified into two main categories: vascular tumors like hemangioma and vascular malformations like hamartoma or lymphaticovenous malformations (LVM) formerly known as lymphangioma (1-6). Vascular tumors are characterized by the presence of rapidly growing, well-defined masses of blood vessels, while vascular malformations are typically present at birth and consist of abnormally formed blood vessels that do not proliferate like tumors (6). Beside of these benign tumors and malformations, Kaposis’s sarcoma is a rare malignant tumor (2). All these lesions can affect one or both eyes, and may cause symptoms e.g. blurred vision, foreign body sensation, redness, epiphora and bleeding. Treatment strategies depend on etiology, size and location of the lesion as well as patient’s symptoms, and may include observation, surgical excision, laser- and cryo therapy, injection of therapeutic agents among others. (1-9). We describe a patient with an isolated conjunctival LVM, who presented with cystic hyposphagma to our emergency department.

Purpose: Proliferative vitreoretinopathy (PVR) is currently treated surgically. Reliable pharmaceutical options would be desirable, and numerous drugs have been proposed. This in vitro study is intended to systematically compare and determine the most promising candidates for the treatment of PVR.Methods: A structured literature review was conducted in the "PubMed" database to identify previously published agents proposed for medical treatment of PVR -36 substances that met the inclusion criteria. Toxicity and antiproliferative effects were evaluated on primary human retinal pigment epithelial (hRPE) using colorimetric viability assays. The seven substances with the widest therapeutic range between toxicity and no longer detectable antiproliferative effect were then validated with a bromodeoxyuridine assay and a scratch wound healing assay using primary cells derived from surgically excised human PVR membranes (hPVR).Results: Among 36 substances, 12 showed no effect on hRPE at all. Seventeen substances had a significant (P < 0.05) toxic effect of which nine did not have an antiproliferative effect. Fifteen substances significantly reduced hRPE proliferation (P < 0.05). The seven most promising drugs with the highest difference between toxicity and antiproliferative effects on hRPE were dasatinib, methotrexate, resveratrol, retinoic acid, simvastatin, tacrolimus, and tranilast. Whereof resveratrol, simvastatin, and tranilast additionally showed antiproliferative and dasatinib, resveratrol, and tranilast antimigratory effects on hPVR (P < 0.05).Conclusion: This study presents a systematic comparison of drugs that have been proposed for PVR treatment in a human disease model. Dasatinib, resveratrol, simvastatin, and tranilast seem to be promising and are well-characterized in human use.