Corneal Endothelium Of Patients Research Articles

Corneal Endothelial Morphological Alterations in Type 2 Diabetes: Associations with Glycemic Control, Disease Duration, and Diabetic Retinopathy Severity—A Cross-Sectional Study in Benghazi, Libya

Background The corneal endothelium is essential for maintaining corneal transparency and fluid regulation, and its dysfunction can lead to corneal edema and decreased visual acuity. Corneal specular microscopy is used to detect corneal disorders early. Aim This study aimed to evaluate morphological changes in the corneal endothelium of patients with type 2 diabetes mellitus (DM) using specular microscopy. Methods This study enrolled 50 individuals with type 2 diabetes and 50 nondiabetic individuals as control subjects. Patients with certain ocular and systemic conditions were excluded. The collected data included demographic information, medical history, recent hemoglobin A1c (HbA1c) values, visual acuity, ophthalmological examination, and diabetic retinopathy (DR) staging. The corneal endothelial morphology was evaluated using a noncontact specular microscope, which measured corneal endothelial cell density (ECD), central corneal thickness (CCT), coefficient of variation of cell size (CV), and hexagonality. The data were analyzed using SPSS software, and the results were presented as mean ± standard deviation, numbers, and percentages. An unpaired t-test was used to compare different means, and a p-value less than 0.05 was considered statistically significant. Results A majority of diabetic patients (62%) had diabetes for more than 10 years, and 58% of them had higher than 7.5% HbA1c levels. In terms of DR severity, 46% of patients had mild nonproliferative diabetic retinopathy (NPDR), 36% had moderate NPDR, and 10% had severe NPDR. There were significant differences between diabetic and nondiabetic groups in corneal ECD (2480 ± 223 cells/mm2 for DM group vs. 2652 ± 234 cells/mm2 for non-DM), hexagonality (39.6 ± 2.8% for DM group vs. 47 ± 2.1% for non-DM), CV (42 ± 2.9% for DM group vs. 35.5 ± 2.3% for non-DM), and CCT (550 ± 14.8 µm for DM group vs. 530 ± 9.6 µm for non-DM). Patients who had diabetes for more than 10 years had significantly lower ECD (2356 cells/mm2 vs. 2689 cells/mm2), lower hexagonality (39 vs. 41%), and higher CV (43 vs. 41%) and higher CCT (553.9 ± 4.6 vs. 545.5 ± 4.0) than those with less than 10 years of diabetes. As the severity of DR increased, there was a significant decrease in ECD (from 2641 ± 194 cells/mm2 for mild NPDR to 2310 ± 82 cells/mm2 for severe PDR), a decrease in hexagonality (from 40.9 ± 3.2% for mild PDR to 37.4 ± 1.1% for severe PDR), an increase in CV (from 40.1 ± 2.3% for mild PDR to 44 ± 2.9% for severe PDR), and an increase in CCT (from 543.7 ± 13.8 for mild PDR 563.8 ± 2.9 for severe PDR). Conclusion This study revealed that type 2 diabetes is associated with significant reductions in ECD and hexagonality, as well as an increase in CV and CCT. The severity of DR and the duration of diabetes were correlated with changes in these parameters. These findings underscore the need for a comprehensive evaluation of corneal health in diabetic patients.

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.

Evaluation of corneal endothelium and correlation with disease severity in patients with Graves’ ophthalmopathy: A specular microscopy-based study

PurposeTo investigate the possible corneal endothelial damage in Graves’ ophthalmopathy (GO) and its relationship with GO activity. MethodsThis cross-sectional study included 101 eyes of 55 patients with GO. Each eye was assigned a specific clinical activity score (CAS). Accordingly, they were classified as active (CAS ≥ 3) or inactive (CAS< 3). The corneal endothelium was measured using a non-contact specular microscope (Tomey EM-4000; Tomey Corp.). Endothelial cell density (ECD), average cell area (ACA), standard deviation of cell area (SD), coefficient of variation in cell area (CV), hexagonal cell ratio (HEX), and central corneal thickness (CCT) were recorded. ResultsAmong the eyes included in the study, 71 had inactive GO and 30 had active GO. ACA and HEX levels were lower (p<0,001) and CV values were higher (p<0.001) in patients with GO than in healthy subjects. Corneal endothelial cell morphology was altered in active GO compared to inactive GO. The SD (p = 0,009) and CV (p<0,001) were significantly higher in active GO than in inactive GO. When the parameters examined were correlated with CAS, a statistically significant positive correlation was observed between proptosis (p = 0,036, r = 0,385) and CV (p = 0,001, r = 0,595). ConclusionOur study confirmed that morphological changes occur in the corneal endothelium of patients with GO. CV and SD values, in conjunction with CAS, can be used as non-invasive and quantitative indices to examine the activity status of GO. The demonstration of endothelial changes even in GO eyes with low CAS may be considered an incentive to include non-contact specular microscopy in the routine clinical evaluation of all patients with GO.

Gene Expression and Missplicing in the Corneal Endothelium of Patients With a TCF4 Trinucleotide Repeat Expansion Without Fuchs' Endothelial Corneal Dystrophy.

PurposeCTG trinucleotide repeat (TNR) expansion in an intron of the TCF4 gene is the most common genetic variant associated with Fuchs' endothelial corneal dystrophy (FECD). Although several mechanisms have been implicated in the disease process, their exact pathophysiologic importance is unclear. To understand events leading from TCF4 TNR expansion to disease phenotype, we characterized splicing, gene expression, and exon sequence changes in a rare cohort of patients with TNR expansions but no phenotypic FECD (RE+/FECD−).MethodsCorneal endothelium and blood were collected from patients undergoing endothelial keratoplasty for non-FECD corneal edema. Total RNA was isolated from corneal endothelial tissue (n = 3) and used for RNASeq. Gene splicing and expression was assessed by Mixture of Isoforms (MISO) and MAP-RSeq software. Genomic DNA was isolated from blood mononuclear cells and used for whole genome exome sequencing. Base calling was performed using Illumina's Real-Time Analysis.ResultsThree genes (MBNL1, KIF13A, AKAP13) that were previously identified as misspliced in patients with a CTG TNR expansion and FECD disease (RE+/FECD+) were found normally spliced in RE+/FECD− samples. Gene expression differences in pathways associated with the innate immune response, cell signaling (e.g., TGFβ, WNT), and cell senescence markers were also identified between RE+/FECD− and RE+/FECD+ groups. No consistent genetic variants were identified in RE+/FECD− patient exomes.ConclusionsIdentification of novel splicing patterns and differential gene expression in RE+/FECD− samples provides new insights and more relevant gene targets that may be protective against FECD disease in vulnerable patients with TCF4 CTG TNR expansions.

Effect of Trinucleotide Repeat Expansion on the Expression of TCF4 mRNA in Fuchs' Endothelial Corneal Dystrophy.

PurposeCTG trinucleotide repeat (TNR) expansion is frequently found in transcription factor 4 (TCF4) in Fuchs' endothelial corneal dystrophy (FECD), though the effect of TNR expansion on FECD pathophysiology remains unclear. The purpose of this study was to evaluate the effect of TNR expansion on TCF4 expression in corneal endothelium of patients with FECD.MethodsPeripheral blood DNA and Descemet membrane with corneal endothelium were obtained from 203 German patients with FECD. The CTG TNR repeat length in TCF4 was determined by short tandem repeat (STR) assays and Southern blotting using genomic DNA. Genotyping of rs613872 in TCF4 was performed by PCR. TCF4 mRNA levels in corneal endothelium were evaluated by quantitative PCR using three different probes. Control corneal endothelial samples were obtained from 35 non-FECD subjects.ResultsThe STR assay and Southern blotting showed that 162 of the 203 patients with FECD (80%) harbored CTG trinucleotide repeat lengths larger than 50. Quantitative PCR using all three probes demonstrated that TCF4 mRNA is significantly upregulated in the corneal endothelium of patients with FECD, regardless of the presence of TNR expansion. However, the length of the TNR tended to show a positive correlation with TCF4 expression level. No correlation was shown between the genotype of TCF4 SNP, rs613872, and the level of TCF4 expression.ConclusionsOur findings showed that TCF4 mRNA is upregulated in the corneal endothelium of patients with FECD. Further studies on the effects of TCF4 upregulation on corneal endothelial cell function will aid in understanding the pathophysiology of FECD.

Repeat-Associated Non-ATG (RAN) Translation in Fuchs' Endothelial Corneal Dystrophy.

PurposeThe strongest genetic association with Fuchs' endothelial corneal dystrophy (FECD) is the presence of an intronic (CTG·CAG)n trinucleotide repeat (TNR) expansion in the transcription factor 4 (TCF4) gene. Repeat-associated non-ATG (RAN) translation, an unconventional protein translation mechanism that does not require an initiating ATG, has been described in many TNR expansion diseases, including myotonic dystrophy type 1 (DM1). Given the similarities between DM1 and FECD, we wished to determine whether RAN translation occurs in FECD.MethodsAntibodies against peptides in the C-terminus of putative RAN translation products from TCF4 were raised and validated by Western blotting and immunofluorescence (IF). CTG·CAG repeats of various lengths in the context of the TCF4 gene were cloned in frame with a 3× FLAG tag and transfected in human cells. IF with antipeptide and anti-FLAG antibodies, as well as cytotoxicity and cell proliferation assays, were performed in these transfected cells. Corneal endothelium derived from patients with FECD was probed with validated antibodies by IF.ResultsCTG·CAG repeats in the context of the TCF4 gene are transcribed and translated via non-ATG initiation in transfected cells and confer toxicity to an immortalized corneal endothelial cell line. An antipeptide antibody raised against the C-terminus of the TCF4 poly-cysteine frame recognized RAN translation products by IF in cells transfected with CTG·CAG repeats and in FECD corneal endothelium.ConclusionsExpanded CTG·CAG repeats in the context of the third intron of TCF4 are transcribed and translated via non-ATG initiation, providing evidence for RAN translation in corneal endothelium of patients with FECD.

Activation of TGF-\u03b2 signaling induces cell death via the unfolded protein response in Fuchs endothelial corneal dystrophy

Fuchs endothelial corneal dystrophy (FECD) is a slowly progressive bilateral disease of corneal endothelium in which accumulation of extracellular matrix (ECM) and loss of corneal endothelial cells (CECs) are phenotypic features. The corneal endothelium maintains corneal transparency by regulating water hydration; consequently, corneal endothelial dysfunction causes serious vision loss. The only therapy for corneal haziness due to corneal endothelial diseases, including FECD, is corneal transplantation using donor corneas, and no pharmaceutical treatment is available. We provide evidence that the expression levels of transforming growth factor-β (TGF-β) isoforms and TGF-β receptors are high in the corneal endothelium of patients with FECD. A cell model based on patients with FECD shows that TGF-β signaling induced a chronic overload of ECM proteins to the endoplasmic reticulum (ER), thereby enhancing the formation of unfolded protein and triggering the intrinsic apoptotic pathway through the unfolded protein response (UPR). We propose that inhibition of TGF-β signaling may represent a novel therapeutic target that suppresses cell loss as well as the accumulation of ECM in FECD.

Identification of Circulating Fibrocytes and Dendritic Derivatives in Corneal Endothelium of Patients With Fuchs' Dystrophy.

Fuchs' endothelial corneal dystrophy (FECD) is a degenerative eye disorder affecting 4% of Americans older than 40. It is the leading indication for corneal endothelial (CE) transplantation for which there is a global donor shortage. This study aimed to gain further insight into the pathophysiology of FECD and identify targets for nonsurgical therapy. CE from patients with late-onset FECD was compared with that of normal controls using microarray expression analysis (n = 4 FECD, n = 4 normal), reverse transcriptase quantitative PCR (n = 9 FECD, n = 8 normal), and immunohistology (n = 55 FECD, n = 15 normal). This led to the identification of circulating fibrocytes and their dendritic derivatives in all examined CE samples with FECD (in all clinical stages of symptomatic FECD and independent of prior cataract surgery). These cells were not present in normal CE. In this study we characterize their morphology, protein expression profile, number, and localization within the CE layer of patients with FECD. Circulating fibrocytes and their dendritic derivatives are a new aspect of FECD that deserves further investigation. Because they are known to cause fibrosis in a variety of organs, they may play a similar role in FECD and might be a valuable target for nonsurgical therapy.

Immune Cells on the Corneal Endothelium of an Allogeneic Corneal Transplantation Rabbit Model.

Corneal endothelial cell density undergoes a progressive decrease for many years after transplantation, eventually threatening patients with late endothelial failure. The purpose of this study was to investigate the possibility of an immunologic response in successfully grafted corneal endothelium. The corneal endothelium of patients who had undergone corneal transplantation was evaluated by specular microscopy. Rabbit models were subjected to penetrating keratoplasty (PK) with either syngeneic or allogeneic corneal transplants and Descemet's stripping endothelial keratoplasty (DSEK) with allogeneic corneal transplants. The presence of immune cells and expression of proinflammatory cytokines were determined by immunostaining. The corneal endothelium and immune cells were also evaluated by scanning electron microscopy. Scanning slit contact specular microscopy of patients with no features of graft rejection revealed cell-like white dots on the grafted corneal endothelium. The corneal endothelium of the allogeneic PK and DSEK rabbit models displayed the presence of immune cells, including CD4+ T-helper cells, CD8+ cytotoxic T cells, CD20+ B lymphocytes, CD68+ macrophages, and neutrophils, but these immune cells were rarely observed in the syngeneic PK model. These immune cells also produced proinflammatory cytokines. Notably, some of the corneal endothelial cells situated near these immune cells exhibited features of apoptosis. T lymphocytes, B lymphocytes, macrophages, and neutrophils are present on the grafted corneal endothelium in both PK and DSEK allogeneic rabbit models. The potential involvement of immune cells as an underlying pathophysiology for late endothelial failure deserves further examination.

The analysis of corneal endothelium of patients with cataract

Objective To investigate the corneal endothelial in cataract patients combined with different diseases and evaluate the endurance of cornea in cataract surgery.Methods 277 eyes of 198 patients were divided into seven groups based on accompanied eye disease.The corneal endothelial cell density (CD),average cell area (AVE) and percentage of hexagonal cell density(HEXA)were detected in each eye.The data were analyzed by one-way analysis of variance with SPSS 16.0.Results There were 7 groups:group A (senile cataract),group B1 (cataract combined with acute primary angle closure glaucoma),group B2(cataract combined with chronic primary angle closure glaucoma),group C1 (cataract combined with non-proliferative diabetic retinopathy),group C2 (cataract combined with proliferative diabetic retinopathy),group D (cataract combined with uveitis),group E (cataract combined with high myopia).Compared with group A,CD was lower and AVE was higher in group B1 and C2 significantly (P ＜ 0.05).HEXA was lower in group C2 than group A.Compared with group A,there were no significant differences in CD,AVE and HEXA in group B2,C1,D and E (P ＞ 0.05).Conclusion Acute primary angle closure glaucoma and proliferative diabetic retinopathy have influence on corneal endothelia; chronic primary angle closure glaucoma,non-proliferative diabetic retinopathy,uveitis and high myopia have no obvious influence on corneal endothelia. Key words: Endothelia, corneal ; Glaucoma ; Diabetes ; Uveitis ; Myopia, high

Biomechanical profile of the cornea in primary congenital glaucoma

The aim of our study was to investigate the biomechanical properties of the cornea in primary congenital glaucoma (PCG) and to identify the potential ocular determinants, which affect the corneal biomechanical metrics. Corneal hysteresis (CH), corneal resistance factor (CRF) and central corneal thickness (CCT) were measured in 26 patients with PCG (40 eyes) with the aid of ocular response analyser. In vivo laser-scanning confocal microscopy was used for the estimation of stromal keratocyte density (KD) and the evaluation of corneal endothelium. Twenty normal subjects (40 eyes) served as controls. Student's t-test and Pearson's correlation coefficients were used for statistical analysis. p Values <0.05 were considered statistically significant. Corneal hysteresis, CRF and CCT were significantly reduced in patients with PCG (all p < 0.05). Corneal hysteresis and CRF negatively correlated with the corneal diameter in both groups (r(1) = -0.53, r(2) = -0.66, p < 0.001 for CH and r(1) = -0.61, r(2) = -0.69, p < 0.001 for CRF). Moreover, we identified a significant correlation between CH and CRF with CCT in both groups (r(1) = 0.51, r(2) = 0.48, p < 0.001 for CH and r(1) = 0.45, r(2) = 0.44, p < 0.001 for CRF). Mean KD was significantly reduced both in the anterior and posterior corneal stroma in patients with PCG (764 ± 162 and 362 ± 112 cells/mm(2) , respectively) compared with controls (979 ± 208 and 581 ± 131 cells/mm(2) , respectively) (p < 0.001). There was no significant correlation between the keratocyte density in anterior and/or posterior stroma and CH or CRF in any group (r(1) = 0.29, r(2) = 0.31, p < 0.06). Mean endothelial cell density was also significantly reduced in PCG group (2920 ± 443 cells/mm(2) ) compared with control group (3421 ± 360 cells/mm(2) ) (p < 0.001). Pleomorphism and polymegalism were significantly increased in corneal endothelium of patients with PCG. Our results showed a significant reduction in CH and CRF in PCG. Both CH and CRF were negatively correlated with corneal diameter. A significant correlation of CH and CRF with CCT was identified in both groups. Keratocyte density was decreased in PCG, but did not have a significant impact on CH and CRF. Mean endothelial density was also decreased in PCG. Our results suggest that reduced CCT and increased corneal diameter are major ocular determinants for the modified corneal biomechanical profile in PCG, while cellular alterations in corneal stroma and endothelium have no significant biomechanical impact.

Demonstration of “Owl’s Eye” Morphology by Confocal Microscopy in a Patient With Presumed Cytomegalovirus Corneal Endotheliitis

To report confocal microscopic observations of characteristic corneal endothelial lesions in a patient with presumed cytomegalovirus (CMV) corneal endotheliitis. Case report. A 77-year-old, immunocompetent man was admitted with corneal edema, keratic precipitates, and coin-shaped lesions in the right eye. Confocal microscopy was performed to examine the corneal endothelium. Polymerase chain reaction (PCR) was used to identify viral DNA in an aqueous humor sample. CMV DNA was detected by PCR. Confocal microscopy showed large corneal endothelial cells with an area of high reflection in the nucleus surrounded by a halo of low reflection. This "owl's eye" morphology is characteristic of CMV infection. Topical and intravenous ganciclovir treatment resulted in rapid resolution of the corneal precipitates and edema, followed by disappearance of the owl's eye morphology. Confocal microscopy can detect the owl's eye morphology in the corneal endothelium of patients with presumed CMV corneal endotheliitis.

Corneal endothelial changes in patients with chronic renal failure

PURPOSE: To evaluate the morphologic changes of the corneal endothelium in patients with chronic renal failure. METHODS: Twenty corneas of 20 patients with chronic renal failure undergoing hemodialysis were examined by wide-field specular microscopy. Twenty normal corneas from 20 healthy age-matched subjects were enrolled as control corneas. RESULTS: Despite normal endothelial cell density, the corneal endothelium of patients with chronic renal failure showed marked polymegethism and pleomorphism. CONCLUSION: Polymegethism and pleomorphism of the corneal endothelium are found in patients with chronic renal failure.