Analysis Of Corneal Endothelial Cells Research Articles

Comparison of automated corneal endothelial cell analysis in healthy and postoperative eyes with phakic intraocular lens: a cross-sectional study and literature review

PurposeMeasurement of corneal endothelial cells is critical for postoperative evaluation of phakic intraocular lens (pIOL) surgery. However, inter-instrument differences in corneal endothelial cell density (ECD) after pIOL implantation have not yet been reported. This study aimed to compare automated corneal endothelial cell analysis between CellChek-20 (Konan Medical, Hyogo, Japan) and EM-4000 (Tomey, Nagoya, Japan) in healthy and postoperative eyes with pIOL.MethodsWe retrospectively analyzed 154 healthy and 236 postoperative eyes after pIOL surgery. Endothelial cell measurements were performed using CellChek-20 and EM-4000 with autofocusing and automated image analysis. ECD, percentage of hexagonal cells (HEX), coefficient of variation in cell size (CoV), and central corneal thickness (CCT) were compared between the two devices.ResultsThe ECDs of the two devices were highly correlated in both healthy (Spearman’s correlation coefficient [r] = 0.805; p < 0.001) and postoperative (r = 0.901; p < 0.001) groups. ECD from CellChek-20 was higher than EM-4000 in both healthy (mean difference = 228.9 cells/mm2; p < 0.001) and postoperative (mean difference = 115.6 cells/mm2; p < 0.001) groups. The CCT values also showed a strong correlation in healthy eyes (r = 0.974; p < 0.001) and in postoperative eyes (r = 0.936; p < 0.001); however, significant inter-instrument differences were observed. HEX and CV showed significant differences and relatively weak correlations (r < 0.7) between the two devices in both healthy and postoperative groups.ConclusionThe ECD values between the two instruments were correlated, but that of the CellChek-20 was significantly higher than that of the EM-4000 in both healthy and postoperative eyes after pIOL surgery. Most previous studies have also shown that the Konan software overestimated the ECD compared to other products in automatic measurement mode. The possibility of measurement bias should be considered when replacing equipment used for corneal endothelial cell measurements.

Canine corneal endothelial cell analysis using vital dyes and light microscopy.

To evaluate the use of vital dyes and light microscopy for assessing canine corneal endothelial morphology exvivo. The corneas of 40 canine eyes (n = 20 dogs) enucleated <24 h following euthanasia or death were isolated and flat-mounted on a slide. Corneal endothelium was stained via 0.25% trypan blue followed by 0.5% alizarin red (pH 4.2), photographed, then the following morphological features were calculated using ImageJ: mean cell density (MCD), mean cell area (MCA), polymegathism (coefficient of variation of cell area), and pleomorphism (% hexagonality). Mean ± standard deviation (range) outcomes were: MCD, 2544 ± 541 cells/mm2 (1750-3922 cells/mm2); MCA, 431 ± 97 μm2 (251-626 μm2); polymegathism, 17 ± 2% (14%-22%); pleomorphism, 84 ± 3% (80%-90%). No significant differences (p ≥ .122) were noted for any outcome between male versus female or brachycephalic versus non-brachycephalic dogs. Young dogs (<10 years) had lower MCA (p = .044), lower pleomorphism (p = .003), and higher MCD (p = .035) when compared to older dogs (≥10 years). Age was significantly (p ≤ .049) correlated with MCA (r = 0.467), MCD (r = -0.476), polymegathism (r = 0.444), and pleomorphism (r = 0.609). The combination of vital dyes and light microscopy allowed for clear visualization and evaluation of the corneal endothelium in canine eyes exvivo. Our findings can be used in future studies to deepen our understanding of the corneal endothelium in health and disease.

Integrated Analysis of Transcriptome and Proteome of the Human Cornea and Aqueous Humor Reveal Novel Biomarkers for Corneal Endothelial Cell Dysfunction.

Earlier studies have reported that elevated protein levels in the aqueous humor (AH) are associated with corneal endothelial cell dysfunction (CECD), but the details of the underlying mechanism as well as specific biomarkers for CECD remain elusive. In the present study, we aimed to identify protein markers in AH directly associated with changes to corneal endothelial cells (CECs), as AH can be easily obtained for analysis. We carried out an in-depth proteomic analysis of patient-derived AH as well as transcriptomic analysis of CECs from the same patients with bullous keratopathy (BK) resulting from CECD. We first determined differentially expressed genes (DEGs) and differentially expressed proteins (DEPs) from CECs and AH in CECD, respectively. By combining transcriptomic and proteomic analyses, 13 shared upregulated markers and 22 shared downregulated markers were observed between DEGs and DEPs. Among these 35 candidates from biomarker profiling, three upregulated markers were finally verified via data-independent acquisition (DIA) proteomic analysis using additional individual AH samples, namely metallopeptidase inhibitor 1 (TIMP1), Fc fragment of IgG binding protein (FCGBP), and angiopoietin-related protein 7 (ANGPTL7). Furthermore, we confirmed these AH biomarkers for CECD using individual immunoassay validation. Conclusively, our findings may provide valuable insights into the disease process and identify biofluid markers for the assessment of CEC function during BK development.

Analysis of Corneal Endothelial Cells Measured by Noncontact Specular Microscopy in 20s

Analysis of Corneal Endothelial Cells Measured by Noncontact Specular Microscopy in 20s

RNA-Seq–based transcriptome analysis of corneal endothelial cells derived from patients with Fuchs endothelial corneal dystrophy

Fuchs endothelial corneal dystrophy (FECD) is the most common inherited corneal disease. Fibrillar focal excrescences called guttae and corneal edema due to corneal endothelial cell death result in progressive vision loss. Multiple genetic variants have been reported, but the pathogenesis of FECD is not fully understood. In this study, we used RNA-Seq to analyze differential gene expression in the corneal endothelium obtained from patients with FECD. Differential expression analysis of transcriptomic profiles revealed that expression of 2366 genes (1092 upregulated and 1274 downregulated genes) was significantly altered in the corneal endothelium of patients with FECD compared to healthy subjects. Gene ontology analysis demonstrated an enrichment of genes involved in extracellular matrix (ECM) organization, response to oxidative stress, and apoptotic signaling. Several pathway analyses consistently indicated the dysregulation of ECM-associated pathways. Our differential gene expression findings support the previously proposed underlying mechanisms, including oxidative stress and apoptosis of endothelial cells, as well as the phenotypic clinical FECD hallmark of ECM deposits. Further investigation focusing on differentially expressed genes related to these pathways might be beneficial for elucidating mechanisms and developing novel therapies.

U-Net Convolutional Neural Network for Segmenting the Corneal Endothelium in a Mouse Model of Fuchs Endothelial Corneal Dystrophy.

The purpose of this study was to assess the U-Net-based convolutional neural network performance for segmenting corneal endothelium and guttae of Fuchs endothelial corneal dystrophy. Twenty-eight images of corneal endothelial cells and guttae of Col8a2L450W/L450W knock-in mice were obtained by specular microscopy. We used 20 images as training data to develop the U-Net for analyzing guttae and cell borders. The proposed network was validated using independent test data of 8 images. Cell density, hexagonality, and coefficient of variation were calculated from the predicted cell borders and compared with ground truth. U-Net allowed the prediction of cell borders and guttae, and overlays of those segmentations on specular microscopy images highly corresponded to ground truth. The average number of guttae per field was 6.25 ± 8.07 for ground truth and 6.25 ± 7.87 when predicted by the network (Pearson correlation coefficient 0.989, P = 3.25 × 10 -6 ). The guttae areas were 1.60% ± 1.79% by manual determination and 1.90% ± 2.02% determined by the network (Pearson correlation coefficient 0.970, P = 6.72 × 10 -5 ). Cell density, hexagonality, and coefficient of variation analyzed by the proposed network for cell borders showed very strong correlations with ground truth (Pearson correlation coefficient 0.989, P = 3.23 × 10 -6 , Pearson correlation coefficient 0.978, P = 2.66 × 10 -5 , and Pearson correlation coefficient 0.936, P = 6.20 × 10 -4 , respectively). We demonstrated proof of concept for application of U-Net for objective analysis of corneal endothelial cells and guttae in Fuchs endothelial corneal dystrophy, based on limited ground truth data.

Corneal endothelial cell analysis using two non-contact specular microscopes in healthy subjects

To compare specular microscopy values obtained using the Perseus (CSO, Italy) and the Nidek CEM-530 (NIDEK Co., Ltd. Japan) specular microscopes. This prospective study used specular microscopy to examine sixty eyes from thirty healthy subjects (29.83±9.41years; range 18-79years). This was done with both the Nidek CEM-530 and the Perseus on three occasions and results were evaluated by one independent observer. Measurement differences between instruments and agreement between devices were determined. The endothelial cell sample was larger with the Perseus than with the CEM-530 (235.92±38.26 vs. 184.38±43.88, respectively) with a statistically significant difference (P=0.001). Mean endothelial cell density (ECD) with the Perseus and CEM-530 was 2692.75±306.66 and 2556.47±257.38cells/mm(2) (P=0.001), respectively; mean coefficient of variation (CV) was 33.43±4.29 and 28.70±3.82 (P=0.001), respectively, and hexagonality 56.66±6.19% and 68.50±3.64 (P=0.001), respectively. The mean of the differences (Perseus minus CEM-530) for ECD was 136.27±106.16 cells/mm(2) (95% CI 108.85-163.70 cells/mm(2)); 4.73±2.70% (95% CI 4.03-5.41) in CV; and -11.83±5.05 for percentage of hexagonal cells. Both instruments showed significant differences in the measurement of all cell forms, predominantly higher values were found with the Perseus vs the CEM-530, except 4-sided cells. Endothelial cell density, coefficient of variation, and percentage of hexagonal cells between the Perseus and CEM-530 differ statistically. This shows that these instruments should not be used interchangeably.

Comparison of Flex-Center, Center, and Corner Methods of Corneal Endothelial Cell Analysis

The center method of corneal endothelial cell analysis is rapid but excludes the outermost digitized cells of a contiguous group from analysis; the flex-center method (Konan, Inc) is a modification that includes analysis of the outermost cells, which is advantageous in images with few cells. In this study, we examined agreement among the flex-center, center, and corner (standard) methods of endothelial analysis. Identical cells in endothelial images of 10 normal corneas and 10 corneas after penetrating keratoplasty (PK) were analyzed by each method. Agreement among methods for endothelial cell density (ECD), coefficient of variation of cell area (CV), and the percentage of hexagonal cells (HEX) was assessed by using a Student-Newman-Keuls procedure. In normal corneas, there were small (clinically insignificant) differences among methods for ECD (P < 0.001) (mean +/- SD: flex center, 2846 +/- 248 cells/mm; center, 2870 +/- 253 cells/mm; and corner, 2892 +/- 254 cells/mm), CV (P < 0.001) (flex center, 33% +/- 3%; center, 30% +/- 3%; and corner, 30% +/- 3%), and HEX (P = 0.004) (flex center, 60% +/- 6%; center, 60% +/- 8%; and corner, 58% +/- 7%). In PK corneas, the methods agreed for ECD (P = 0.06) (flex center, 908 +/- 319 cells/mm; center, 912 +/- 307 cells/mm; and corner, 929 +/- 333 cells/mm) but disagreed for CV (P = 0.02) (flex center, 35% +/- 13%; center, 30% +/- 10%; and corner, 35% +/- 15%) and HEX (P = 0.02) (flex center, 56% +/- 19%; center, 54% +/- 17%; and corner, 43% +/- 23%). ECD agreed among methods in normal and PK corneas, whereas morphometric data agreed poorly in PK corneas.

Corneal Bee Sting-Induced Endothelial Changes

To report the acute management and clinical findings of a case of corneal bee sting and to report the outcome of corneal endothelial cell analysis 1 year after trauma. Clinical findings, anterior segment photographs, corneal endothelial images, and medical treatment of a case of right corneal bee sting are presented. Right and left central corneal endothelial cell analysis was performed by noncontact specular microscopy. The stinger was removed from the cornea. Systemic, subconjunctival, and topical steroids and systemic and topical antibiotics were given. One year later, a corneal scar and anterior capsular opacity of the lens in the right eye were shown by slit-lamp examination. Endothelial cell analysis determined that the endothelial cell density of the right eye was substantially decreased compared with the left eye. Corneal infiltration gradually decreased, presumably because of the systemic, topical, and subconjunctival steroids. Late complications observed in this case included a substantial decrease in cornea endothelial cell density, a corneal scar, and anterior capsular opacity.

Twenty-three Cases of Primary Cornea Guttata

Purpose: An evaluation of the clinical aspects of patients with primary cornea guttata and the morphology of their corneal endothelial cells. Methods: Twenty-three patients who visited Kanazawa Medical University Hospital or related hospitals in Ishikawa or Fukui prefectures and were diagnosed as having primary cornea guttata by slit lamp microscopy and contact specular microscopy underwent an analysis of corneal endothelial cells as well as their clinical status. Results: In 23 cases, the number of men and young people under 30 years of age was low, with 4 and 2 cases respectively. There was no visual impairment due to cornea guttata. In the specular microscopic findings, the size and density of dark areas varied according to each case and there was asymmetry in some cases. The parameters of the endothelial cells, comprising mean cell area, hexagonality, and the coefficient of variation, were almost within normal limits except for one eye of a 79-year-old man who was speculated to be at an early stage of Fuch's endothelial corneal dystrophy. Three cases had received cataract surgery, but showed no statistical changes in endothelial morphology before or after the surgery. Conclusion: Cornea guttata is thought to be a disease which is not uncommon in Japanese people and which does not bring about visual impairment or abnormalities in corneal endothelial cell parameters except for a small number of patients. Such cases with remarkably decreased cell densities should be considered to be at an early stage of Fuch's dystrophy.

Clinical and specular micrographic findings in corneal guttata and Fuchs' corneal endothelial dystrophy

Focal excrescenses of altered basement membrane material, known as corneal guttata, are a common occurrence in the elderly. A small proportion of those who develop this condition exhibit signs of corneal endothelial dysfunction and progress to Fuchs' corneal endothelial dystrophy. The relationship between the distribution and relative density of guttata and the morphological appearance of the corneal endothelium is as yet poorly understood.In this study, 40 eyes of 20 caucasian patients (5 male and 15 female) identified as having central corneal endothelial guttata were examined (mean age 76 years). Ten eyes noted as having a previous history of ocular trauma, surgery or anterior segment inflammation were excluded from the study. Clinical and specular micrographic findings are compared.Clinical examination confirmed that guttata were most densely distributed in the central cornea. Examination of all four mid‐peripheral quadrants identified the superior cornea as being least severely affected in the majority of cases. In 26 of the 30 eyes examined there was evidence of pigmentary dusting of the corneal endothelium. The severity of guttata did not appear to correlate with the density or distribution of the pigment. In cases of moderate to advanced disease, where guttata had become confluent and thus obscured the view of the corneal endothelium, the clinical differentiation of pigment, guttata and diffuse scarring became difficult. Specular microscopy confirmed the accuracy of the clinical five‐point scale used to quantify the distribution and density of guttata in the study. Quantitative analysis of corneal endothelial cells from the mid‐peripheral cornea indicated that the severity of guttata did not necessarily correlate with a reduced cell count. Central cell counts proved elusive as reflections from the irregular surface created by confluent guttata failed to produce specular images of the underlying endothelium.The authors conclude that the specular microscope is a useful diagnostic tool in the differentiation of guttata from other abnormalities of the endothelium and Descemet's membrane. Specular micrographic imaging also enables the examiner to quantify the distribution and density of guttata. The relationship between the severity of corneal guttata, endothelial cell counts and pigmentation is not well defined and further study of the progression of the disease process is required.

Cartographic system for spatial distribution analysis of corneal endothelial cells.

A combined cartographic and morphometric endothelium analyser has been developed by integrating the HISTO 2000 histological imaging and analysis system with a prototype human corneal endothelium analyser. The complete system allows the elaboration and analysis of cartographies of corneal endothelial tissue, and hence the in vitro study of the spatial distribution of corneal endothelial cells, according to their regional morphometric characteristics (cell size and polygonality). The global cartographic reconstruction is obtained by sequential integration of the data analysed for each microscopic field. Subsequently, the location of each microscopically analysed field is referred to its real position on the histologic preparation by means of X-Y co-ordinates; both are provided by micrometric optoelectronic sensors installed on the optical microscope stage. Some cartographies of an excised human corneal keratoconus button in vitro are also presented. These cartographic images allow a macroscopic view of endothelial cells analysed microscopically. Parametric colour images show the spatial distribution of endothelial cells, according to their specific morphometric parameters, and exhibit the variability in size and cellular shape which depend on the analysed area.