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Didar S Anwar, Matthew M Kruger, V Vinod Mootha Department of Ophthalmology, University of Texas Southwestern Medical Center, Dallas, TX, USA Objective: We describe a modified technique for performing lamellar dissection in deep anterior lamellar keratoplasty after failure to achieve a “big bubble” detachment of Descemet’s membrane (DM) with deep intrastromal pneumatic injection. Methods: The technique utilizes blunt lamellar dissection with blunt-tipped corneal mini scissors as an alternative to a crescent blade, which can be difficult for surgeons to master and is associated with a high risk of perforation. Results: Other techniques of blunt dissection, such as the Melles technique, cannot be utilized after failure to achieve a big bubble, as emphysema in the stroma prevents visualization of the spatula. In contrast, our blunt scissors lamellar dissection technique takes advantage of the emphysema and microdetachments of DM created by the pneumatic injection. Conclusion: This technique provides deep anterior lamellar keratoplasty surgeons with a simple, alternative method of baring DM or achieving a pre-DM plane with minimal residual stroma, after failure to achieve a big bubble. Keywords: scissors, blunt, dissection, big bubble, DALK

Purpose: The purpose of this study was to determine the prevalence of glaucoma and/or ocular hypertension (G/OHTN) in patients with Fuchs endothelial corneal dystrophy (FECD) and correlate with FECD severity and TCF4 cytosine-thymine-guanine18.1 (CTG18.1) trinucleotide repeat expansion genotype., Methods: We included 167 FECD probands and 110 controls from the University of Texas Southwestern Medical Center FECD Genetics Study to estimate the association between FECD and G/OHTN. Participants underwent slit-lamp microscopy for the assessment of Krachmer grade disease severity of FECD. The diagnosis of G/OHTN was ascertained using a patient-reported history of G/OHTN, previous glaucoma surgery and/or glaucoma laser procedure, and use of glaucoma drops. Genomic DNA from blood of participants was used to genotype the CTG18.1 repeat polymorphism by fragment analysis using short tandem repeat and triplet repeat primed polymerase chain reaction assays., Results: We observed a 19.2% prevalence of G/OHTN in the FECD probands compared with that of 7.3% in controls. The odds ratio of developing G/OHTN in FECD cases compared with controls was estimated to be 3.34 with a 95% confidence interval of 1.42-7.79 adjusting for age and sex. Among FECD cases, the likelihood of developing G/OHTN correlated positively with Krachmer grade (P = 0.043) and age (P = 0.026). There was no statistical difference of the proportions of patients developing G/OHTN between FECD cases with and without TCF4 CTG18.1 repeat expansion (16 out of 94 and 15 out of 72, respectively, P > 0.05)., Conclusions: Patients with clinically significant FECD should be routinely monitored for the development of glaucoma regardless of their TCF4 repeat expansion genotype., Competing Interests: V. V. Mootha is coinventor on patent application related to treatments and biomarkers for Fuchs' dystrophy. The remaining authors have no conflicts of interest to disclose., (Copyright © 2024 The Author(s). Published by Wolters Kluwer Health, Inc.)

Background: Late-onset Fuchs' endothelial corneal dystrophy (FECD) is a degenerative disease of cornea and the leading indication for corneal transplantation. Genetically, FECD patients can be categorized as with (RE+) or without (RE-) the CTG trinucleotide repeat expansion in the transcription factor 4 gene. The molecular mechanisms underlying FECD remain unclear, though there are plausible pathogenic models proposed for RE+ FECD., Method: In this study, we performed a meta-analysis on RNA sequencing datasets of FECD corneal endothelium including 3 RE+ datasets and 2 RE- datasets, aiming to compare the transcriptomic profiles of RE+ and RE- FECD. Gene differential expression analysis, co-expression networks analysis, and pathway analysis were conducted., Results: There was a striking similarity between RE+ and RE- transcriptomes. There were 1,184 genes significantly upregulated and 1,018 genes significantly downregulated in both RE+ and RE- cases. Pathway analysis identified multiple biological processes significantly enriched in both-mitochondrial functions, energy-related processes, ER-nucleus signaling pathway, demethylation, and RNA splicing were negatively enriched, whereas small GTPase mediated signaling, actin-filament processes, extracellular matrix organization, stem cell differentiation, and neutrophil mediated immunity were positively enriched. The translational initiation process was downregulated in the RE+ transcriptomes. Gene co-expression analysis identified modules with relatively distinct biological processes enriched including downregulation of mitochondrial respiratory chain complex assembly. The majority of oxidative phosphorylation (OXPHOS) subunit genes, as well as their upstream regulator gene estrogen-related receptor alpha (ESRRA), encoding ERRα, were downregulated in both RE+ and RE- cases, and the expression level of ESRRA was correlated with that of OXPHOS subunit genes., Conclusion: Meta-analysis increased the power of detecting differentially expressed genes. Integrating differential expression analysis with co-expression analysis helped understand the underlying molecular mechanisms. FECD RE+ and RE- transcriptomic profiles are much alike with the hallmark of downregulation of genes in pathways related to ERRα-mediated OXPHOS., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2023 Zhang et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

One advantage of antisense oligonucleotides (ASOs) for drug development is their long-lasting gene knockdown after administration in vivo . In this study, we examine the effect on gene expression after intraocular injection in target tissues in the eye. We examined expression levels of the Malat1 gene after intracameral or intravitreal (IV) injection of an anti- Malat1 ASO in corneal epithelium/stroma, corneal endothelium, lens capsule epithelium, neurosensory retina, and retinal pigment epithelium/choroid of the mouse eye. We assessed potency of the compound at 7 days as well as duration of the gene knockdown at 14, 28, 60, 90, and 120 days. The ASO was more potent when delivered by IV injection relative to intracameral injection, regardless of whether the tissues analyzed were at the front or back of the eye. For corneal endothelium, inhibition was >50% after 120 days for ASO at 50 μg. At IV dosages of 6 μg, we observed >75% inhibition of gene expression in the retina and lens epithelium for up to 120 days. ASOs have potential as long-lasting gene knockdown agents in the mouse eye, but efficacy varies depending on the specific ocular target tissue and injection protocol.

Fuchs' corneal endothelial dystrophy (FECD) is a major cause of vision loss. Corneal transplantation is the only effective curative treatment, but this surgery has limitations. A pharmacological intervention would complement surgery and be beneficial for many patients. FECD is caused by an expanded CUG repeat within intron 2 of the TCF4 RNA. Agents that recognize the expanded repeat can reverse the splicing defects associated with the disease. Successful drug development will require diverse strategies for optimizing the efficacy of anti-CUG oligomers. In this study, we evaluate anti-CUG morpholinos conjugated to cyclic cell penetrating peptides. The morpholino domain of the conjugate is complementary to the repeat, while the peptide has been optimized for import across cell membranes. We show that morpholino conjugates can enter corneal endothelial cells and block the CUG RNA foci associated with the disease. These experiments support morpholino peptide conjugates as an approach for developing anti-CUG therapies for FECD., Competing Interests: The authors declare the following competing financial interest(s): X.S., M.K., M.D.S., and Z.Q. are employees of Entrada Therapeutics., (© 2023 The Authors. Published by American Chemical Society.)

Purpose: In the United States, 70% of Fuchs' endothelial corneal dystrophy (FECD) cases are caused by an intronic trinucleotide repeat expansion in the TCF4 gene. CUG repeat RNA transcripts from this expansion accumulate as nuclear foci in the corneal endothelium. In this study, we sought to detect foci in other anterior segment cell types and assess their molecular impact., Methods: We examined CUG repeat RNA foci appearance, expression of downstream affected genes, gene splicing, and TCF4 RNA expression in corneal endothelium, corneal stromal keratocytes, corneal epithelium, trabecular meshwork cells, and lens epithelium., Results: CUG repeat RNA foci, the hallmark of FECD in corneal endothelium (found in 84% of endothelial cells), are less detectable in trabecular meshwork cells (41%), much less prevalent in stromal keratocytes (11%) or corneal epithelium (4%), and absent in lens epithelium. With few exceptions including mis-splicing in the trabecular meshwork, differential gene expression and splicing changes associated with the expanded repeat in corneal endothelial cells are not observed in other cell types. Expression of the TCF4 transcripts including full-length isoforms containing the repeat sequence at the 5' end is much higher in the corneal endothelium or trabecular meshwork than in the corneal stroma or corneal epithelium., Conclusions: Expression of the CUG repeat containing TCF4 transcripts is higher in the corneal endothelium, likely contributing to foci formation and the large molecular and pathologic impact on those cells. Further studies are warranted to examine any glaucoma risk and impact of the observed foci in the trabecular meshwork of these patients.

Purpose: We sought to define the role of Wwtr1 in murine ocular structure and function and determine the role of mechanotransduction in Fuchs' endothelial corneal dystrophy (FECD), with emphasis on interactions between corneal endothelial cells (CEnCs) and Descemet's membrane (DM)., Methods: A Wwtr1 deficient mouse colony was established, and advanced ocular imaging, atomic force microscope (AFM), and histology/immunofluorescence were performed. Corneal endothelial wound healing was assessed using cryoinjury and phototherapeutic keratectomy in Wwtr1 deficient mice. Expression of WWTR1/TAZ was determined in the corneal endothelium from normal and FECD-affected patients; WWTR1 was screened for coding sequence variants in this FECD cohort., Results: Mice deficient in Wwtr1 had reduced CEnC density, abnormal CEnC morphology, softer DM, and thinner corneas versus wildtype controls by 2 months of age. Additionally, CEnCs had altered expression and localization of Na/K-ATPase and ZO-1. Further, Wwtr1 deficient mice had impaired CEnC wound healing. The WWTR1 transcript was highly expressed in healthy human CEnCs comparable to other genes implicated in FECD pathogenesis. Although WWTR1 mRNA expression was comparable between healthy and FECD-affected patients, WWTR1/TAZ protein concentrations were higher and localized to the nucleus surrounding guttae. No genetic associations were found in WWTR1 and FECD in a patient cohort compared to controls., Conclusions: There are common phenotypic abnormalities seen between Wwtr1 deficient and FECD-affected patients, suggesting that Wwtr1 deficient mice could function as a murine model of late-onset FECD. Despite the lack of a genetic association between FECD and WWTR1, aberrant WWTR1/TAZ protein subcellular localization and degradation may play critical roles in the pathogenesis of FECD.

Objective: Seventy percent of Fuchs' endothelial corneal dystrophy (FECD) cases are caused by an intronic trinucleotide repeat expansion in the transcription factor 4 gene ( TCF4 ). The objective of this study was to characterize the corneal subbasal nerve plexus and corneal haze in patients with FECD with (RE+) and without the trinucleotide repeat expansion (RE-) and to assess the correlation of these parameters with disease severity., Design: Cross-sectional, single-center study., Participants: Fifty-two eyes of 29 subjects with a modified Krachmer grade of FECD severity from 1 to 6 were included in the study. Fifteen of the 29 subjects carried an expanded TCF4 allele length of ≥ 40 cytosine-thymine-guanine repeats (RE+)., Main Outcomes Measures: In vivo confocal microscopy assessments of corneal nerve fiber length (CNFL), corneal nerve branch density, corneal nerve fiber density (CNFD), and anterior corneal stromal backscatter (haze); Scheimpflug tomography densitometry measurements of haze in anterior, central, and posterior corneal layers., Results: Using confocal microscopy, we detected a negative correlation between FECD severity and both CNFL and CNFD in the eyes of RE+ subjects (Spearman ρ = -0.45, P  = 0.029 and ρ = -0.62, P  = 0.0015, respectively) but not in the eyes of RE- subjects. Additionally, CNFD negatively correlated with the repeat length of the expanded allele in the RE+ subjects (Spearman ρ = -0.42, P  = 0.038). We found a positive correlation between anterior stromal backscatter and severity in both the RE+ and RE- groups (ρ = 0.60, P  = 0.0023 and ρ = 0.44, P  = 0.024, respectively). The anterior, central, and posterior Scheimpflug densitometry measurements also positively correlated with severity in both the RE+ and RE- groups ( P  = 5.5 × 10 -5 , 2.5 × 10 -4 , and 2.9 × 10 -4 , respectively, after adjusting for the expansion status in a pooled analysis. However, for patients with severe FECD (Krachmer grades 5 and 6), the posterior densitometry measurements were higher in the RE+ group than in the RE- group ( P < 0.05)., Conclusions: Loss of corneal nerves in FECD supports the classification of the TCF4 trinucleotide repeat expansion disorder as a neurodegenerative disease. Haze in the anterior, central, and posterior cornea correlate with severity, irrespective of the genotype. Quantitative assessments of corneal nerves and corneal haze may be useful to gauge and monitor FECD disease severity in RE+ patients., (© 2022 by the American Academy of Ophthalmology.)

Corneal endothelial dystrophies are a heterogeneous group of diseases with different modes of inheritance and genetic basis for each dystrophy. The genes associated with these diseases encode transcription factors, structural components of the stroma and Descemet membrane, cell transport proteins, and others. Congenital hereditary endothelial dystrophy (CHED) is associated with mutations in two genes, OVOL2 and SLC4A11, for dominant and recessive forms of CHED, respectively. Mutations in three genes are known to cause posterior polymorphous corneal dystrophy (PPCD). They are OVOL2 (PPCD1), ZEB1 (PPCD3), and GRHL1 (PPCD4). The PPCD2 locus involving the collagen gene COL8A2 on chromosome 1 is disputed due to insufficient evidence. Mutations in the COL8A2 gene are associated with early-onset Fuchs' endothelial corneal dystrophy (FECD). Several genes have been associated with the more common, late-onset FECD. Alterations in each of these genes occur in a fraction of patients, and the most prevalent genetic alteration in FECD patients across the world is a triplet repeat expansion in the TCF4 gene. Knowledge of the genetics of corneal endothelial dystrophies has considerably advanced within the last decade and has contributed to better diagnosis of these dystrophies as well as opened up the possibility of novel therapeutic approaches based on the molecular mechanisms involved. The functions of genes identified to date provide insights into the pathogenic mechanisms involved in each disorder., Competing Interests: None

Purpose: Retinitis pigmentosa (RP) is an inherited retinal disorder that results in the degeneration of photoreceptor cells, ultimately leading to severe visual impairment. We characterized a consanguineous family from Southern India wherein a 25 year old individual presented with night blindness since childhood. The purpose of this study was to identify the causative mutation for RP in this individual as well as characterize how the mutation may ultimately affect protein function., Methods: We performed a complete ophthalmologic examination of the proband followed by exome sequencing. The likely causative mutation was identified and modeled in cultured cells, evaluating its expression, solubility (both with western blotting), subcellular distribution, (confocal microscopy), and testing whether this variant induced endoplasmic reticulum (ER) stress (quantitative PCR [qPCR] and western blotting)., Results: The proband presented with generalized and parafoveal retinal pigmented epithelium (RPE) atrophy with bone spicule-like pigmentation in the midperiphery and arteriolar attenuation. Optical coherence tomography scans through the macula of both eyes showed atrophy of the outer retinal layers with loss of the ellipsoid zone, whereas the systemic examination of this individual was normal. The proband's parents and sibling were asymptomatic and had normal funduscopic examinations. We discovered a novel homozygous p.Pro388Ser mutation in the tubby-like protein 1 ( TULP1 ) gene in the individual with RP. In cultured cells, the P388S mutation does not alter the subcellular distribution of TULP1 or induce ER stress when compared to wild-type TULP1, but instead significantly lowers protein stability as indicated with steady-state and cycloheximide-chase experiments., Conclusions: These results add to the list of known mutations in TULP1 identified in individuals with RP and suggest a possible unique pathogenic mechanism in TULP1 -induced RP, which may be shared among select mutations in TULP1 ., (Copyright © 2021 Molecular Vision.)

Purpose: To investigate the long-term corneal changes in patients with Fuchs endothelial corneal dystrophy contributing to superior postoperative visual outcomes after Descemet membrane endothelial keratoplasty (DMEK) compared with Descemet stripping automated endothelial keratoplasty (DSAEK)., Methods: Using retrospective analysis, we evaluated 9 patients with Fuchs endothelial corneal dystrophy who underwent DSAEK in 1 eye and DMEK in the fellow eye. Patients were genotyped for the triplet repeat expansion in the TCF4 gene and imaged using optical coherence tomography, Scheimpflug imaging, and in vivo confocal microscopy through focusing., Results: Eight of 9 subjects were genotyped, and all were found to harbor the triplet repeat expansion. The average time between endothelial keratoplasty and imaging was 76 ± 22 and 37 ± 9 months after DSAEK and DMEK, respectively. The mean best spectacle-corrected visual acuity (logMAR) was 0.04 ± 0.05 and 0.11 ± 0.03 in the DMEK eyes versus DSAEK eyes (P = 0.02), respectively. Posterior corneal higher order aberrations were less in the DMEK eyes compared with fellow DSAEK eyes (0.25 ± 0.06 and 0.66 ± 0.25, respectively, P ≤ 0.01). Using confocal microscopy through focusing, we found that the persistent anterior stromal haze was correlated between the right and left eyes (R = 0.73, P ≤ 0.05), but total stromal backscattering was higher for the DSAEK eyes (P ≤ 0.05)., Conclusions: DSAEK inherently results in higher total stromal backscattering (haze) compared with DMEK because of the addition of stromal tissue. Lower higher order aberrations of the posterior cornea and lower total stromal backscattering (haze) may both contribute to superior visual outcomes after DMEK compared with DSAEK.

Purpose: Fuchs' endothelial corneal dystrophy (FECD) is the leading indication for corneal transplantation. Seventy percent of cases are caused by an intronic CTG triplet repeat expansion in the TCF4 gene that results in accumulation of pathogenic expanded CUG repeat RNA (CUG exp ) as nuclear foci in corneal endothelium. A catalytically dead Cas9 (dCas9) can serve as an effective guide to target genomic DNA or RNA transcripts. Here, we examined the utility of the clustered regularly interspaced short palindromic repeats (CRISPR)-dCas9 system to effectively target and reduce CUG exp ., Methods: We delivered dCas9 and repeat-targeting single guide RNA (sgRNA) expression plasmids to patient-derived endothelial cells using lipofection or lentiviral transduction. We used fluorescence in situ hybridization (FISH) and RNA dot-blot hybridization to quantify CUG exp foci and repeat RNA levels, respectively. TCF4 expression levels were assessed using quantitative PCR (qPCR)., Results: Using FISH, we found that expression of both dCas9 and a (CAG) n sgRNA complementary to CUG exp are necessary to reduce foci. We observed a reduction in percentage of cells with foci from 59% to 5.6% and number of foci per 100 cells from 73.4 to 7.45 ( P < 0.001) in cells stably expressing dCas9-(CAG) n sgRNA but saw no decrease in cells expressing dCas9-(CUG) n sgRNA or nontargeting control sgRNA. In cells with dCas9-(CAG) n sgRNA, we detected a reduction in CUG exp RNA by dot-blot without any reduction in TCF4 mRNA levels using qPCR., Conclusions: Using CRISPR-dCas9 to target the trinucleotide repeat is a promising treatment for FECD contingent on effective in vivo delivery., Translational Relevance: This work advances a gene therapy for a common age-related degenerative disorder., Competing Interests: Disclosure: Z. Rong, None; X. Gong, None; J.D. Hulleman, None; D.R. Corey, None; V.V. Mootha (P), (Copyright 2020 The Authors.)

Antisense oligonucleotides (ASOs) are synthetic nucleic acids that recognize complementary RNA sequences inside cells and modulate gene expression. In this study, we explore the feasibility of ASO delivery to the cornea. We used quantitative polymerase chain reaction to test the efficacy of a benchmark ASO targeting a noncoding nuclear RNA, Metastasis-Associated Lung Adenocarcinoma Transcript 1 ( MALAT1 ), in a human corneal endothelial cell line, ex vivo human corneas, and in vivo in mice. In vivo delivery was via intravitreal or intracameral injections as well as topical administration. The anti- MALAT1 ASO significantly reduced expression of MALAT1 in a corneal endothelial cell line. We achieved a dose-dependent reduction of target gene expression in endothelial tissue from ex vivo human donor corneas. In vivo mouse experiments confirmed MALAT1 reduction in whole corneal tissue via intravitreal and intracameral routes, 82% and 71% knockdown, respectively ( P  < 0.001). Effects persisted up to at least 21 days, 32% ( P  < 0.05) and 43% ( P  < 0.05) knockdown, respectively. We developed protocols for the isolation and analysis of mouse corneal endothelium and observed reduction in MALAT1 expression upon both intravitreal and intracameral administrations, 64% ( P  < 0.05) and 63% ( P  < 0.05) knockdown, respectively. These data open the possibility of using ASOs to treat corneal disease.

How genetic defects trigger the molecular changes that cause late-onset disease is important for understanding disease progression and therapeutic development. Fuchs' endothelial corneal dystrophy (FECD) is an RNA-mediated disease caused by a trinucleotide CTG expansion in an intron within the TCF4 gene. The mutant intronic CUG RNA is present at one-two copies per cell, posing a challenge to understand how a rare RNA can cause disease. Late-onset FECD is a uniquely advantageous model for studying how RNA triggers disease because: (i) Affected tissue is routinely removed during surgery; (ii) The expanded CTG mutation is one of the most prevalent disease-causing mutations, making it possible to obtain pre-symptomatic tissue from eye bank donors to probe how gene expression changes precede disease; and (iii) The affected tissue is a homogeneous single cell monolayer, facilitating accurate transcriptome analysis. Here, we use RNA sequencing (RNAseq) to compare tissue from individuals who are pre-symptomatic (Pre&#95;S) to tissue from patients with late stage FECD (FECD&#95;REP). The abundance of mutant repeat intronic RNA in Pre&#95;S and FECD&#95;REP tissue is elevated due to increased half-life in a corneal cells. In Pre&#95;S tissue, changes in splicing and extracellular matrix gene expression foreshadow the changes observed in advanced disease and predict the activation of the fibrosis pathway and immune system seen in late-stage patients. The absolute magnitude of splicing changes is similar in pre-symptomatic and late stage tissue. Our data identify gene candidates for early drivers of disease and biomarkers that may represent diagnostic and therapeutic targets for FECD. We conclude that changes in alternative splicing and gene expression are observable decades prior to the diagnosis of late-onset trinucleotide repeat disease., (© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Purpose: Fuchs' endothelial corneal dystrophy (FECD) is a major cause of vision loss and the most common nucleotide repeat disorder, affecting 4% of United States population greater than 40 years of age. Seventy percent of FECD cases are due to an intronic CTG expansion within the TCF4 gene, resulting in accumulation of CUG repeat RNA nuclear foci in corneal endothelium. Each endothelial cell has approximately two sense foci, and each focus is a single RNA molecule. This study aimed to obtain a better understanding of how rare repeat RNA species lead to disease., Methods: We quantitatively examined muscleblind-like (MBNL) proteins and their interaction with foci in both patient-derived corneal endothelial cell lines and human corneal endothelial tissue., Results: Using fluorescent in situ hybridization and immunofluorescence, we found that depletion of both MBNL1 and MBNL2 reduces nuclear RNA foci formed by the repeat, suggesting that both are necessary for foci. Quantitative studies of RNA and protein copy number revealed MBNLs to be abundant in the total cellular pool in endothelial cell lines but are much lower in human corneal endothelial tissue. Studies using human tissue nuclear and cytoplasmic fractions indicate that most MBNL proteins are localized to the cytoplasm., Conclusions: The low levels of MBNL1/2 in corneal tissue, in combination with the small fraction of protein in the nucleus, may make corneal endothelial cells especially susceptible to sequestration of MBNL1/2 by CUG repeat RNA. These observations may explain how a limited number of RNA molecules can cause widespread alteration of splicing and late-onset degenerative FECD.

Fuchs' endothelial corneal dystrophy (FECD) leads to vision loss and is one of the most common inherited eye diseases. Corneal transplants are the only curative treatment available, and there is a major unmet need for treatments that are less invasive and independent of donor tissue. Most cases of FECD are associated with an expanded CUG repeat within the intronic region of TCF4 and the mutant RNA has been implicated as the cause of the disease. We previously presented preliminary data suggesting that single-stranded antisense oligonucleotides (ASOs) can inhibit CUG RNA foci in patient-derived cells and tissue. We now show that duplex RNAs and single-stranded silencing RNAs (ss-siRNAs) reduce the number of cells with foci and the number of foci per cells. Potencies are similar to those that are achieved with chemically modified ASOs designed to block foci. These data widen the potential for synthetic nucleic acids to be used to treat a widely prevalent and debilitating disease.

Purpose: To determine if CTG18.1 TNR expansion length prognosticates the clinical progression of Fuchs' Endothelial Corneal Dystrophy (FECD)., Methods: This was a prospective cohort study. A total of 51 patients with newly diagnosed FECD were recruited and followed-up over a period of 12 years, from November 2004 to April 2016. Baseline clinical measurements included central corneal thickness (CCT), endothelial cell density (ECD) and CTG18.1 TNR expansion length from peripheral leukocytes, with yearly repeat measurements of CCT and ECD. A patient was defined to have experienced significant clinical progression and to have developed Threshold Disease if any of these criteria were fulfilled in either eye: a) CCT increased to >700μm, b) ECD decreased to <700 cells/mm2, or c) underwent keratoplasty for treatment of FECD., Results: Patients were categorized as having at least one allele whose maximum allele length was equal to or greater than 40 repeats (L≥40, n = 22, 43.1%), or having both alleles shorter than 40 repeats (L<40). Threshold Disease rates at the 5-year time point were 87.5% for the L≥40 group and 47.8% for the L<40 group (p = 0.012). This difference narrowed and was no longer statistically significant at the 8-years (92.9% vs 78.9%, p = 0.278) and 10-years (92.9% vs 84.2%, p = 0.426) time points., Conclusions: L≥40 patients are at greater risk of FECD progression and development of Threshold Disease within the first 5 years following diagnosis., Competing Interests: The authors have declared that no competing interests exist.

Purpose: Fuchs' endothelial corneal dystrophy (FECD) caused by the CTG triplet repeat expansion in the TCF4 gene (CTG18.1 locus) is the most common repeat expansion disorder. Intergenerational instability of expanded repeats and clinical anticipation are hallmarks of other repeat expansion disorders. In this study, we examine stability of triplet repeat allele length and FECD disease severity in parent-child transmission of the expanded CTG18.1 allele., Methods: We studied 44 parent-child transmissions of the mutant expanded CTG18.1 allele from 26 FECD families. The CTG18.1 polymorphism was genotyped using short tandem repeat analysis, triplet repeat primed PCR assay, and Southern blot analysis. FECD severity was assessed using modified Krachmer grading (KG) system. Triplet repeat length of mutant allele and KG severity were compared between generations., Results: Instability of the expanded allele was seen in 14 of 44 (31.8%) parent-child transmissions, and the likelihood of an unstable event increased with the size of the parental allele (P = 5.9 x 10^-3). A tendency for contraction was seen in transmission of large alleles (repeat length > 120), whereas intermediate alleles (repeat length between 77 and 120) had predilection for further expansion (P = 1.3 x 10^ - 3). Although we noted increased KG severity in the offspring in three pairs, none of these transmissions were associated with allele instability., Conclusions: We observed instability of the TCF4 triplet repeat expansion in nearly a third of parent-child transmissions. Large mutant CTG18.1 alleles are prone to contraction, whereas intermediate mutant alleles tend to expand when unstably transmitted. Intergenerational instability of TCF4 repeat expansion has implications on FECD disease inheritance.

Fuchs' endothelial corneal dystrophy (FECD) is the most common repeat expansion disorder. FECD impacts 4% of U.S. population and is the leading indication for corneal transplantation. Most cases are caused by an expanded intronic CUG tract in the TCF4 gene that forms nuclear foci, sequesters splicing factors and impairs splicing. We investigated the sense and antisense RNA landscape at the FECD gene and find that the sense-expanded repeat transcript is the predominant species in patient corneas. In patient tissue, sense foci number were negatively correlated with age and showed no correlation with sex. Each endothelial cell has ∼2 sense foci and each foci is single RNA molecule. We designed antisense oligonucleotides (ASOs) to target the mutant-repetitive RNA and demonstrated potent inhibition of foci in patient-derived cells. Ex vivo treatment of FECD human corneas effectively inhibits foci and reverses pathological changes in splicing. FECD has the potential to be a model for treating many trinucleotide repeat diseases and targeting the TCF4 expansion with ASOs represents a promising therapeutic strategy to prevent and treat FECD.

Purpose: To examine the outcomes of resident-performed manual small-incision cataract surgery (SICS) in an urban academic setting., Setting: Parkland Memorial Hospital, Dallas, Texas, USA., Design: Retrospective case series., Methods: Manual SICS was used only in selected cases for which phacoemulsification was expected to be difficult, namely for mature or brunescent cataracts, traumatic cataracts, and pseudoexfoliation syndrome or other causes of zonular weakness. All manual SICS cases performed by resident physicians as the primary surgeon over a 5-year period were reviewed. Postoperative visual acuity, intraoperative complications, and early postoperative complications were the main outcomes measured., Results: For the 52 cases identified, the mean preoperative visual acuity was 2.165 logarithm of the minimum angle of resolution (logMAR) ± 0.141 (SD) (95% confidence interval) (slightly better than had motion acuity), improving to 0.278 ± 0.131 logMAR (Snellen 20/38) corrected visual acuity postoperatively. Of the 52 cases, the most frequent intraoperative complications were iris prolapse (5 cases [9.6%]) and zonular dialysis (4 cases [7.7%]), with vitreous loss occurring in 1 case (1.9%). The most frequent postoperative complications were cystoid macular edema (3 cases [5.8%]), retained ophthalmic viscosurgical device (2 cases [3.8%]), intraocular lens displacement (2 cases [3.8%]), and microhyphema (2 cases [3.8%])., Conclusions: Although the more advanced wound construction in manual SICS might be challenging to surgeons unfamiliar with the technique, it was a safe and efficacious technique in the hands of learning residents. With several advantages over phacoemulsification, such as cost and ability to remove very dense nuclei, manual SICS will play a valuable role in modern cataract surgery., (Copyright © 2018. Published by Elsevier Inc.)

Purpose: The most common cause of Fuchs' endothelial corneal dystrophy (FECD) is an intronic CTG repeat expansion in TCF4. Expanded CUG repeat RNA colocalize with splicing factor, muscleblind-like 1 (MBNL1), in nuclear foci in endothelium as a molecular hallmark. Myotonic dystrophy type 1 (DM1) is a neuromuscular disorder caused by a CTG repeat expansion in the 3'-untranslated region (UTR) of DMPK. In this study, we examine for RNA-MBNL1 foci in endothelial cells of FECD subjects with DM1, test the hypothesis that DM1 patients are at risk for FECD, and determine prevalence of TCF4 and DMPK expansions in a FECD cohort., Methods: Using FISH, we examined for nuclear RNA-MBNL1 foci in endothelial cells from FECD subjects with DM1. We examined 13 consecutive unrelated DM1 patients for FECD using slit-lamp and specular microscopy. We genotyped TCF4 and DMPK repeat polymorphisms in a FECD cohort of 317 probands using short-tandem repeat and triplet repeat-primed PCR assays., Results: We detected abundant nuclear RNA foci colocalizing with MBNL1 in endothelial cells of FECD subjects with DM1. Six of thirteen DM1 patients (46%) had slit-lamp and specular microscopic findings of FECD, compared to 4% disease prevalence (P = 5.5 × 10-6). As expected, 222 out of 317 (70%) FECD probands harbored TCF4 expansion, while one subject harbored DMPK expansion without prior diagnosis of DM1., Conclusions: Our work suggests that DM1 patients are at risk for FECD. DMPK mutations contribute to the genetic burden of FECD but are uncommon. We establish a connection between two repeat expansion disorders converging upon RNA-MBNL1 foci and FECD.

The structure of the cornea is vital to its transparency, and dystrophies that disrupt corneal organization are highly heritable. To understand the genetic aetiology of Fuchs endothelial corneal dystrophy (FECD), the most prevalent corneal disorder requiring transplantation, we conducted a genome-wide association study (GWAS) on 1,404 FECD cases and 2,564 controls of European ancestry, followed by replication and meta-analysis, for a total of 2,075 cases and 3,342 controls. We identify three novel loci meeting genome-wide significance (P<5 × 10 -8 ): KANK4 rs79742895, LAMC1 rs3768617 and LINC00970/ATP1B1 rs1200114. We also observe an overwhelming effect of the established TCF4 locus. Interestingly, we detect differential sex-specific association at LAMC1, with greater risk in women, and TCF4, with greater risk in men. Combining GWAS results with biological evidence we expand the knowledge of common FECD loci from one to four, and provide a deeper understanding of the underlying pathogenic basis of FECD.

Purpose: To describe 2 cases of congenital corneal endothelial edema resulting from novel de novo mutations., Methods: Case A patient was a 15-month-old white child and case B patient was a 3-year-old Hispanic child presenting with bilateral cloudy corneas since birth. Clinicopathologic findings are presented. DNA samples were screened for mutations in candidate genes by Sanger sequencing., Results: Slit-lamp examination of case A patient revealed stromal edema and haze. Histology of the keratoplasty button showed stromal thickening with loss of endothelium and thin Descemet membrane. Sanger sequencing established the diagnosis of congenital hereditary endothelial dystrophy by detection of a compound heterozygous mutation in SLC4A11. The proband displayed a novel de novo frameshift mutation in one SLC4A11 allele, p.(Pro817Argfs*32), in conjunction with a maternally inherited missense mutation in SLC4A11, p.(Arg869His). Case B patient similarly presented with stromal edema and stromal haze. Histopathologic analysis revealed a spongy epithelium, focal discontinuities in Bowman layer, stromal thickening with areas of compacted posterior stroma, variable thickness of Descemet membrane, and regional multilayered endothelium. Sanger sequencing found a novel de novo nonsense mutation in the first exon of ZEB1, p.(Cys7*)., Conclusions: To the authors' knowledge, we report the earliest clinical presentation of posterior polymorphous corneal dystrophy resulting from a de novo mutation in ZEB1. Additionally, we present a congenital hereditary endothelial dystrophy case with a thin Descemet membrane with a novel compound heterozygous SLC4A11 mutation. In the absence of a family history or consanguinity, de novo mutations may result in congenital corneal endothelial dystrophies.

Purpose: To identify the causative mutation in two siblings from a consanguineous family in India with retinitis pigmentosa (RP) and polydactyly without other findings of Bardet-Biedl syndrome (BBS). We also performed functional characterization of the mutant protein to explore its role in this limited form of BBS., Methods: The siblings underwent a thorough ophthalmological examination, including retinal optical coherence tomography (OCT) imaging, and an extensive physical examination with abdominal ultrasonography to characterize the disease phenotype. Next-generation sequencing (NGS) using a panel targeting retinal degeneration genes was performed on genomic DNA samples from the siblings and parents. Upon identification of the causative mutation, functional characterization was accomplished by performing protein-protein interaction studies in human embryonic kidney (HEK-293T) and human adult retinal pigmented epithelium (ARPE-19) cells., Results: The two siblings showed signs of RP and polydactyly. The patients did not have truncal obesity, renal anomalies, hydrometrocolpos, congenital heart disease, or overt cognitive defects. NGS identified a homozygous c.1184A>G mutation in the MKKS/BBS6 gene in both patients resulting in a p.H395R substitution in the MKKS/BBS6 protein. This mutant protein decreased the interaction of MKKS/BBS6 with BBS12 but did so to a different extent in the HEK-293T versus ARPE-19 cells. Nonetheless, the effect of the H395R variant on disrupting interactions with BBS12 was not as profound as other reported MKKS/BBS6 mutations associated with syndromic RP., Conclusions: We identified a novel H395R substitution in MKKS/BBS6 that results in a unique phenotype of only RP and polydactyly. Our observations reaffirm the notion that mutations in MKKS/BBS6 cause phenotypic heterogeneity and do not always result in classic MKKS or BBS findings.

Importance: The CTG18.1 triplet repeat expansion in TCF4 has recently been found to be a common functional variant contributing significant risk to the development of Fuchs endothelial corneal dystrophy (FECD) in Eurasian populations., Objectives: To determine the effect of the expanded CTG18.1 allele of TCF4 on FECD severity and to correlate CTG triplet repeat allele length to the severity of FECD., Design, Setting, and Participants: In a cross-sectional analysis, we studied 139 index cases (probands and unrelated individuals) with FECD recruited from a cornea referral practice at the University of Texas Southwestern Medical Center, Dallas, from April 2010 through February 2015. The triplet repeat polymorphism CTG18.1 was genotyped using a combination of short tandem repeat analysis, triplet repeat-primed polymerase chain reaction assay, and Southern blot analysis. Severity of FECD was graded using a modified Krachmer grading system (severity scale of 0-6 based on extent of confluent guttae)., Main Outcomes and Measures: The CTG triplet repeat length of the largest allele was compared with the Krachmer grade of FECD severity, keratoplasty proportion, and central corneal thickness in the white subset., Results: Eighty-five of 122 white index cases with FECD (69.7%) harbored the triplet repeat expansion. The mean (SD) Krachmer grade was 5.61 (0.76) in the group with the repeat expansion compared with 5.11 (1.05) in the group without the expanded repeats (P = .01). Forty-seven participants with the repeat expansion (55.3%) had undergone keratoplasty at the time of recruitment, compared with 13 (35.1%) of those without the expansion (P = .0497). There was a positive correlation of Krachmer grade to triplet repeat number (P = .002) and a nominal association of the keratoplasty proportion with triplet repeat number (P = .04). The mean (SD) central corneal thickness was 605.9 (50.5) μm in the group with the expanded repeats compared with 581.3 (50.5) μm in the group without the expansion (P = .04)., Conclusions and Relevance: The Krachmer grade of disease severity was greater in FECD cases with the CTG18.1 triplet repeat expansion in TCF4 than in those without the expansion. The CTG triplet repeat allele length was positively correlated with the Krachmer grade of severity. The TCF4 triplet repeat expansion resulted in a more severe form of FECD, with clinical and surgical therapeutic implications.

This review study provides information regarding the construction, design, and use of six commercially available endothelial allograft insertion devices applied for Descemet's stripping automated endothelial keratoplasty (DSAEK). We also highlight issues being faced in DSAEK and discuss the methods through which medical devices such as corneal inserters may alleviate these issues. Inserter selection is of high importance in the DSAEK procedure since overcoming the learning curve associated with the use of an insertion device is a time and energy consuming process. In the present review, allograft insertion devices were compared in terms of design, construction material, insertion technique, dimensions, incision requirements and endothelial cell loss to show their relative merits and capabilities based on available data in the literature. Moreover, the advantages/disadvantages of various insertion devices used for allograft insertion in DSAEK are reviewed and compared. The information presented in this review can be utilized for better selection of an insertion device for DSAEK.

Objective: To correlate corneal endothelium-Descemet membrane (EDM) layer parameters of scroll tightness with donor age, endothelial cell density (ECD), and history of diabetes., Methods: Endothelium-Descemet membrane layer scrolls were harvested from 26 corneoscleral buttons using the SCUBA technique by a cornea-fellowship trained ophthalmologist masked to donor age. Two independent outcome parameters were used to characterize the scrolling severity of successfully harvested tissue: scroll width and tendency for EDM scroll formation (referred to as scroll rating on a 1-4 scale: incomplete scroll formation to tightly scrolled)., Results: Mean donor age was 59 ± 17 (15-69) years. Mean ECD of EDM scroll was 2,451 ± 626 (range: 1,307-3,195) cells per square millimeter. Using stepwise linear regression, a significant correlation was found between scroll width and donor age (R=0.497, P<0.05). Additionally, a significant inverse correlation was found between scroll width and ECD (R=-0.605, P<0.05). There was no statistically significant correlation between a donor history of diabetes and the parameters of scrolling tendency., Conclusions: Our data suggest that using older donors reduces EDM scroll tightness.

Purpose: Expansion of the intronic CTG18.1 triplet repeat locus within TCF4 contributes significant risk to the development of Fuchs' endothelial corneal dystrophy (FECD) in Eurasian populations, but the mechanisms by which the expanded repeats result in degeneration of the endothelium have been hitherto unknown. The purpose of this study was to examine FECD endothelial samples for the presence of RNA nuclear foci, the hallmark of toxic RNA, as well as evidence of haploinsufficiency of TCF4., Methods: Using fluorescence in situ hybridization, we examined for the presence of nuclear RNA foci containing expanded CUG transcripts in corneal endothelial samples from FECD subjects with CTG18.1 expansion. We also examined for any changes in expression levels of TCF4 by quantitative real-time PCR., Results: Numerous discrete nuclear RNA foci were identified in endothelial samples of FECD subjects (n = 8) harboring the CTG18.1 expansion, but not in controls lacking the expansion (n = 5) (P = 7.8 × 10(-4)). Percentage of cells with foci in expansion-positive endothelial samples ranged from 33% to 88%. RNA foci were absent in endothelial samples from an FECD subject without CTG18.1 expansion and a subject with endothelial dysfunction without FECD. Expression of the constitutive TCF4 exon encoding the basic helix-loop-helix domain was unaltered with CTG18.1 expansion., Conclusions: Our findings suggest that the RNA nuclear foci are pathognomonic for CTG18.1 expansion-mediated endothelial disease. The RNA nuclear foci have been previously found only in rare neurodegenerative disorders caused by repeat expansions. Our detection of abundant ribonuclear foci in FECD implicates a role for toxic RNA in this common disease., (Copyright 2015 The Association for Research in Vision and Ophthalmology, Inc.)

Purpose: To test the association between the CTG18.1 trinucleotide repeat expansion of TCF4 gene and Fuchs' endothelial corneal dystrophy (FECD) in a Chinese population., Methods: The trinucleotide repeat polymorphism CTG18.1 was genotyped using short tandem repeat and triplet repeat primed polymerase chain reaction assays in 57 Chinese subjects with FECD and 121 controls. Statistical association of the expanded CTG18.1 allele and 18 single nucleotide polymorphisms (SNPs) across TCF4 with FECD was evaluated. To investigate the linkage disequilibrium structure of the TCF4 region, haplotype analysis was performed on our study subjects and compared with genotyping data of 97 Han Chinese and 85 Caucasians in the 1000 Genomes Project., Results: The expanded CTG18.1 allele was associated with FECD (P = 4.7 × 10(-14)), with the odds ratio of each copy of the expanded allele estimated to be 66.5 (95% confidence interval: 12.6-350.1). Five TCF4 SNPs showed association with FECD at a nominal level (P < 5.0 × 10(-2)); however, conditional on the expanded CTG18.1 polymorphism, none of the SNPs showed association with FECD. The only haplotype associated with the disease was the one with the expansion at the CTG18.1 locus., Conclusions: Transethnic replication of the association between the CTG18.1 repeat expansion in the TCF4 gene and FECD suggests it is a common, causal variant shared in Eurasian populations conferring significant risk for the development of FECD. Our data suggest that the expanded CTG18.1 allele is the main, if not sole, causal variant at this gene locus in the Chinese population., (Copyright 2014 The Association for Research in Vision and Ophthalmology, Inc.)

Purpose: To describe the phenotypic characteristics and clinical course of a sporadic case of congenital fibrosis of the extraocular muscles (CFEOM) and Möbius syndrome with a de novo mutation in the KIF21A gene encoding a kinesin motor protein., Methods: An individual with the rare combination of CFEOM and Möbius syndrome underwent comprehensive ophthalmologic and neurological evaluations. Magnetic resonance imaging (MRI) including diffusion tensor imaging (DTI) tractigraphy at 3T field strength was used to evaluate orbital, encephalic, and intracranial nerve integrity. The proband and her healthy parents underwent screening for mutations in the KIF21A, PHOX2A, and TUBB3 genes., Results: The patient exhibited congenital, nonprogressive, bilateral external ophthalmoplegia, bilateral ptosis, bilateral facial palsy, and developmental delay. Her inability to blink resulted in severe exposure keratopathy and subsequent corneal perforation requiring a penetrating keratoplasty. MRI revealed an unremarkable configuration of the axial central nervous system and preservation of the intracranial portion of cranial nerves I, II, III, V, VI, VII, and VIII (cranial nerve IV is not normally visualized by MRI). A novel and de novo heterozygous KIF21A mutation (c.1056C>G, p.Asp352Glu) in a highly conserved region of the gene was present in the proband., Conclusions: The reported KIF21A D352E mutation and associated phenotype further expand the clinical and mutational spectrum of CFEOM and Möbius syndrome.

Purpose: We tested the association between two intronic polymorphisms (CTG18.1 and rs613872) in TCF4 and Fuchs' endothelial corneal dystrophy (FECD), and analyzed their segregation patterns in families., Methods: We recruited 120 unrelated Caucasian subjects with FECD and 100 controls. Available family members of probands were recruited. Genotyping of the single nucleotide polymorphism (SNP) rs613872 was performed using Sanger sequencing or real-time allelic discrimination assay. The trinucleotide repeat polymorphism, CTG18.1, was genotyped using a combination of short tandem repeat assay and triplet repeat primed PCR assay. The cytosine-thymine-guanine (CTG) repeat length of ≥40 was classified as an expanded CTG18.1 allele. Association of the two loci with FECD was evaluated. Segregation in 29 families was examined., Results: The two polymorphisms are in linkage disequilibrium (r(2) = 0.65 in cases and 0.31 in controls). Significant associations were found between FECD and rs613872 (P = 3.1 × 10(-17)), expanded CTG18.1 allele (P = 6.5 × 10(-25)), and their haplotypes (P = 5.9 × 10(-19)). The odds ratio (OR) of each copy of the rs613872 G allele for FECD was estimated to be 9.5 (95% confidence interval [CI], 5.1-17.5). The OR of each copy of the CTG18.1 expanded allele was estimated to be 32.3 (95% CI, 13.4-77.6). The expanded CTG 18.1 allele cosegregated with the trait in 52% (15/29) of families with complete penetrance and 10% (3/29) with incomplete penetrance., Conclusions: We report, to our knowledge, the first independent replication of the expanded CTG 18.1 allele conferring significant risk for FECD (>30-fold increase). The expanded allele cosegregates with the trait with complete penetrance in a majority of families, but we also document cases of incomplete penetrance.

Purpose: We investigated effects of smoking and other risk factors on the development of advanced Fuchs' endothelial corneal dystrophy (FECD) and on central corneal thickness (CCT)., Methods: Eyes from Caucasian probands, affected and unaffected family members, and unrelated controls matched for age from the FECD Genetics Multi-Center Study (n = 2044 subjects) were examined. Univariate and multivariate models, adjusted for family correlations, were used to determine the effect of smoking, sex, diabetes, and age on FECD case/control status and CCT., Results: In a multivariate model, sex and smoking were associated significantly with advanced FECD (grades 4-6) development (P = 0.016 and P = 0.047, respectively). Female sex increased odds by 34%. Smoking increased odds by 30%. In a multivariate model, diabetes was associated with an increase of 9.1 μm in average CCT (P = 0.021). Female sex was associated significantly with a decrease in average CCT by 6.9 μm (P = 0.015). Smoking had no significant effect on CCT in any model. As shown previously, advanced FECD was associated with large increases in CCT (31.4-94.2 μm)., Conclusions: Smoking was associated with an increased risk of advanced FECD and self-reported diabetes was associated with increased CCT. Further study of the impact of smoking and diabetes on FECD development and changes in corneal thickness is warranted.

Purpose: The purpose of this study was to describe a new modification for big-bubble deep anterior lamellar keratoplasty (DALK) using pneumatic pressure to detach Descemet membrane (DM) via air injection followed by ophthalmic viscoelastic device (OVD) injection., Methods: After failure of big-bubble formation after air injection, OVD was injected from a different site other than the previous air injection using a 27-gauge cannula to detach DM, called air-visco bubble (AVB) DALK technique. The technique was used in 7 human corneoscleral rims that were investigated with anterior segment optical coherence tomography and histopathology and in 69 eyes that underwent DALK surgeries., Results: Big-bubble formation was noted in 4 of 7 of the donor corneoscleral rims. The anterior segment optical coherence tomography showed big-bubble formations together with intrastromal OVD accumulation. The histology of the donor corneas showed microdetachments at the DM in the periphery, deep intrastromal separation, and big-bubble formation filled with OVD. One hundred forty-one of 210 eyes (67%) underwent successful DALK with only air injection, and 69 of 210 eyes (33%) underwent AVB technique when a big bubble was not achieved with only air injection. All the corneas showed a clear interface with good wound healing when DM was bared with the AVB DALK technique., Conclusions: Additional OVD injection to detach DM may be useful in cases where air injection fails. Also, creating small DM detachments with air injection may facilitate the formation of a big bubble with further OVD injection.

Purpose: To describe the first reported case of traumatic globe rupture after femtosecond laser-assisted penetrating keratoplasty., Methods: A 21-year-old Hispanic man suffered a corneal laceration after blunt trauma leading to a visually significant corneal scar and aphakia., Results: The patient underwent femtosecond laser-assisted penetrating keratoplasty using a zigzag-shaped wound with secondary intraocular lens placement. Eighteen months after surgery, he suffered another blunt trauma resulting in inferior dehiscence of his inferior penetrating keratoplasty wound, loss of uveal tissue, and extrusion of the scleral-fixated posterior chamber intraocular lens. The patient underwent repair of the wound dehiscence., Conclusions: Despite the theoretical advantages conferred by a shaped graft-host junction, a shaped penetrating keratoplasty wound created with femtosecond laser can still be subject to dehiscence from trauma.

Purpose: To report a case series of pupil abnormalities consistent with features of Urrets-Zavalia syndrome (UZS) after Descemet stripping automated endothelial keratoplasty (DSAEK) for corneal edema secondary to corneal endothelial cell dysfunction., Methods: Retrospective chart analysis of subjects who developed UZS after DSAEK at the University of Texas Southwestern Medical Center., Results: We present a series of 7 eyes with features consistent with UZS, after undergoing DSAEK. Elevated intraocular pressures (IOP) were noted in the early postoperative period in all cases. Five of 7 had graft dislocation in the postoperative period and required rebubbling or repeat DSAEK to obtain a well-apposed graft. Patients were followed for 3 to 14 months and showed improvement in visual acuity and IOP, but fixed dilated pupils persisted., Conclusion: A fixed irregular or dilated pupil is a rare complication that can be associated with DSAEK surgery. Patients with an elevated IOP and complicated postoperative course seem to be at greater risk for developing iris ischemia and pupil abnormalities consistent with the diagnosis of UZS.

Purpose: To correlate big-bubble deep anterior lamellar keratoplasty findings in donor corneas with anterior segment optical coherence tomography (OCT) and histology., Methods: This research was conducted entirely at the University of Texas Southwestern Medical Center in Dallas, Texas. We performed deep intrastromal air injections in donor corneas on artificial chambers. Surgical patterns (big bubble, intrastromal emphysema, and perforation) were assessed by spectral domain OCT with a handheld probe and histology., Results: Surgical patterns were evaluated by histology using a novel embedding technique. A classic big bubble may be a Descemet membrane (DM) detachment with a few attached stromal fibrils. There were no large intra-DM separations as previously reported. The emphysematous surgical patterns result from intrastromal emphysema, which can be accompanied by microdetachments of DM. We saw indirect OCT signs of big bubble, but scatter from intrastromal emphysema limits deeper imaging., Conclusions: Surgical patterns of big bubble and intrastromal emphysema correlate with characteristic histology findings. Marked scatter on OCT by intrastromal emphysema limits visualization of deeper corneal structures, but the presence of a big bubble may be inferred.

Objectives: The prevalence of primary open-angle glaucoma (POAG) in patients with corneal endothelial dystrophy has not been previously studied. Prevalence of POAG in patients with endothelial dystrophy was compared with that in the general population to determine the presence of a relationship between the diseases., Design: Retrospective case-control study., Methods: A study of the prevalence of POAG in 430 eyes of 215 patients with endothelial dystrophy was conducted. Patients followed for less than 6 months were excluded. Relative risk of POAG was calculated using age- and race-matched control data from the Baltimore Eye Survey and the Los Angeles Latino Eye Survey for comparison. Ocular hypertension (OHT) and secondary glaucoma (SG) rates after penetrating keratoplasty (PK) and Descemet stripping endothelial keratoplasty (DSEK) were separately analyzed., Results: Relative risk of POAG in white, African American, and Hispanic patients with endothelial dystrophy was 0.94, 2.59, and 3.7, respectively (P = 0.89, 95% confidence interval [CI], -0.028 to 0.0289; P = 0.13; 95% CI, 0.011-0.274; P = 0.055; 95% CI, 0.0423-0.356). Relative risk of SG and combined OHT/SG in PK versus DSEK was 4.15 and 1.95 (P < 0.001; 95% CI, 0.0654-0.322; P = 0.005; 95% CI, 0.116-0.332), respectively. No differences in OHT/SG rates were found comparing PK-triple with PK, DSEK-triple with DSEK, and repeat with primary PK or DSEK (P = 0.98; P = 0.62; P = 0.95; P = 0.87), respectively., Conclusions: No increased risk of POAG was found in patients with endothelial dystrophy. Increased prevalence of OHT/SG was shown with PK versus DSEK; possible mechanisms include mechanical closure of Schlemm's canal by running suture and prolonged steroid use.

Purpose: To evaluate the quality of stromal bed and the safety on endothelium in preparation of donor tissue for Descemet stripping automated endothelial keratoplasty in a masked fashion using 2 mechanical microkeratomes and a femtosecond laser., Methods: Deep anterior lamellar dissection was performed on 15 donor corneas. Central endothelial cell density was calculated using specular microscopy before and after the dissection. One cornea from each of 5 donor pairs was cut with the Moria ALTK system with the CBm microkeratome using the 300-μm head and the mate cut with the Horizon disposable 300-μm microkeratome. Five additional donor corneas were cut with the Intralase 60-kHz FS laser. The donor corneas were then bisected with half of the cornea used for Live/Dead assay to study central endothelial viability. The other halves were sent for scanning electron microscopy of the stromal bed. Qualitative surface roughness of the scanning electron microscopy images was graded by 2 masked observers, and quantitative surface roughness was assessed using roughness evaluation software., Results: The Horizon group showed a smoother stromal bed compared with the Moria or Intralase groups by 2 masked observers. However, the Moria group had the smoothest quantitative score of all the groups when assessed by roughness evaluation software. There was no statistically significant difference among the 3 groups in the percentage change in the central endothelial cell density or percentage of viable central endothelium by Live/Dead assay after the dissection., Conclusions: Both mechanical microkeratomes created smoother stromal bed dissections than the femtosecond laser. All systems provided good endothelial cell viability.

Purpose: To evaluate the long-term refractive and visual outcomes of apodized diffractive multifocal intraocular lens (IOL) implantation after myopic laser in situ keratomileusis (LASIK) with or without subsequent excimer laser enhancement., Setting: University of Texas Southwestern Medical Center at Dallas, Texas, USA., Design: Comparative case series., Methods: This study comprised consecutive patients who had implantation of an AcrySof ReSTOR multifocal IOL after previous LASIK. Included were eyes that had refractive surgery enhancement with a Visx Star S4 excimer laser for residual refractive error correction and eyes that had neodymium:YAG (Nd:YAG) capsulotomy., Results: Of the 49 eyes (38 patients), 21 (42.9%) had laser enhancement and 25 (51.0%) had an Nd:YAG capsulotomy. The mean follow-up was 14.2 months ± 7.23 (SD) (range 9 to 29 months). Although the uncorrected distance visual acuity (UDVA) was significantly worse in eyes with enhancement than in those with no enhancement at 1 month and 6 months (P<.05), there was no significant difference in UDVA between the 2 groups at the last follow-up (P>.05). At the last follow-up, 36 (73.5%) of the 49 eyes had a UDVA of 20/25 or better and an uncorrected near visual acuity of Jaeger 1 or better concurrently; 46 eyes (93.9%) were within ±1.00 diopter (D) of emmetropia, and 41 (83.7%) were within ±0.50 D., Conclusion: Apodized diffractive multifocal IOL implantation and excimer laser enhancement in eyes with previous myopic LASIK provided good results., (Copyright © 2010 ASCRS and ESCRS. Published by Elsevier Inc. All rights reserved.)

Objective: To compare the effect on aqueous tear (AT) evaporation rate of Systane and Optive at 30 min postinstillation in patients with dry eye., Methods: In a crossover study of 20 patients with keratoconjunctivitis sicca, the evaporation rate of AT was measured. Evaporometry was used at two relative humidity (RH) ranges of 25% to 35% and 35% to 45%. The measurements were made at baseline (before the instillation of the study agent) and at 30 min after the instillation of 40 μL of either Systane or Optive per randomization assignment per visit with a 1-week interval between visits., Results: No significant effects on AT evaporation rates at both RHs were found between study agents., Conclusions: In our study, neither Systane nor Optive has a significant impact on AT evaporation at 30 min postinstillation in patients with dry eye.

Purpose: To evaluate alterations in corneal power parameters after Descemet-stripping automated endothelial keratoplasty (DSAEK) using rotating Scheimpflug imaging., Setting: University of Texas, Southwestern Medical Center, Dallas, Texas, USA., Methods: Eyes that had DSAEK were evaluated with a rotating Scheimpflug imaging system (Pentacam). The outcome parameters were mean anterior and posterior keratometry (K), mean anterior radius of curvature and posterior radius of curvature, anterior and posterior astigmatism, central corneal thickness (CCT), true net power, corneal volume, keratometric power deviation, and mean zonal-equivalent K readings. These values were compared with those in a control group of age and sex-matched normal eyes., Results: The DSAEK group comprised 32 eyes (28 patients) and the control group, 32 eyes. The mean anterior K, posterior K, and true net power were 42.53 diopters (D) +/- 1.52 (SD), -6.80 +/- 0.55 D, and 40.55 +/- 1.79 D, respectively, in the DSAEK group and 43.60 +/- 1.62 D, -6.20 +/- 0.31 D, and 42.49 +/- 1.57 D, respectively, in the control group. The posterior radius of curvature, anterior and posterior K values, and true net power were statistically significantly lower in DSAEK eyes than in control eyes (P<.01). The equivalent K readings were significantly lower in all zones in DSAEK eyes than in control eyes (P<.01). The mean CCT was significantly higher in DSAEK eyes than in control eyes (628 +/- 46 mum versus 553 +/- 35 mum) (P<.01)., Conclusion: The true net power of the cornea decreased significantly after DSAEK, mainly because of an increase in posterior curvature., Financial Disclosure: No author has a financial or proprietary interest in any material or method mentioned., (Copyright 2010 ASCRS and ESCRS. Published by Elsevier Inc. All rights reserved.)

Purpose: To identify the incidence of and risk factors for intraocular pressure (IOP) elevation after Descemet stripping automated endothelial keratoplasty (DSAEK)., Methods: Retrospective review was conducted of 68 consecutive DSAEK procedures alone, or in combination with phacoemulsification with intraocular lens implantation or exchange, performed by two surgeons at the University of Texas Southwestern Medical Center between 2005 and 2009. Eyes that developed IOP elevation above 21 mm Hg after DSAEK and requiring initiation or escalation of glaucoma therapy were evaluated., Results: Thirty-seven (54%) eyes showed IOP elevation responsive to medical treatment by a mean follow-up of 11.38 +/- 7.81 months. Six (8.8%) eyes required glaucoma surgery. In the eyes, which developed elevated IOP, gonioscopy did not reveal any new peripheral anterior synechiae formation. Prolonged topical steroid usage, rebubbling, combined DSAEK/cataract surgery, or repeat DSAEK were not significant factors (P>0.05) for development of elevated IOP, but history of previous glaucoma or ocular hypertension (OHTN) was significant (P=0.007)., Conclusions: Intraocular pressure elevation is not uncommon in eyes after DSAEK, but most cases can be controlled with conservative management. Intraocular pressure elevation post-DSAEK occurred by mechanisms other than peripheral anterior synechial angle closure. The only significant risk factor for development of elevated IOP in our series was a previous history of glaucoma or OHTN.

Purpose: To compare the corneal higher-order aberration (HOA) after Descemet's stripping automated endothelial keratoplasty (DSAEK) and penetrating keratoplasty (PKP), and in age-matched controls., Design: Cross-sectional, non-comparative interventional case series., Participants: Thirty-one eyes of 28 patients who underwent DSAEK, 20 eyes of 16 patients who underwent PKP, and 31 eyes of 31 control patients., Intervention: The corneal topography and HOAs of the central 4- and 6-mm zones from anterior and posterior corneal surfaces were evaluated postoperatively with the Scheimpflug rotating imaging system (Oculus Gmbh, Wetzlar, Germany)., Main Outcome Measures: Anterior and posterior corneal HOAs., Results: The mean anterior corneal total HOAs of the central 4 and 6 mm were 0.599+/-0.288 microm and 1.215+/-0.496 microm, respectively, in eyes that underwent DSAEK; 1.730+/-0.826 microm and 3.349+/-1.490 microm, respectively, in eyes that underwent PKP; and 0.439+/-0.163 microm and 0.921+/-0.300 microm, respectively, in controls. Although the mean anterior corneal total HOAs of the central 4 and 6 mm were significantly higher in eyes that underwent PKP than in eyes that underwent DSAEK and in controls (P<0.01), there was no significant difference in anterior corneal total HOAs of the central 4 and 6 mm between eyes that underwent DSAEK and controls. The mean posterior corneal total HOAs of the central 4 and 6 mm were 3.680+/-1.586 microm and 7.142+/-3.011 microm, respectively, in eyes that underwent DSAEK; 2.957+/-1.238 microm and 5.314+/-2.095 microm, respectively, in eyes that underwent PKP; and 0.818+/-0.193 microm and 1.609+/-0.344 microm, respectively, in controls. Although there was no significant difference in posterior corneal total HOAs of the central 4 mm between the DSAEK group and the PKP group, the posterior corneal HOAs of the central 6 mm were significantly higher in the DSAEK group than in the PKP group (P<0.01)., Conclusions: Although posterior corneal HOAs are significantly higher in eyes that underwent DSAEK, anterior corneal HOAs are not significantly different in eyes that underwent DSAEK than those of age-matched controls., (Copyright 2010 American Academy of Ophthalmology. Published by Elsevier Inc. All rights reserved.)

Purpose: To evaluate the efficacy, predictability, safety, and intraoperative and postoperative complications of laser in situ keratomileusis (LASIK) retreatment in myopic eyes using wavefront-guided ablation with iris registration (IR)., Methods: Retrospective analysis was used to evaluate wavefront-guided retreatment with IR in a consecutive cohort of 77 eyes (57 patients) after LASIK. The eyes were divided into two groups: no previous retreatment group (group 1) (n = 63) and previous LASIK retreatment group (group 2) (n = 14). The primary outcome variables assessed postoperatively at 1, 3, and 6 months were uncorrected visual acuity (UCVA), best-corrected visual acuity (BCVA), and pre- and postretreatment changes in manifest refraction., Results: The mean preretreatment spherical equivalent in group 1 was reduced from -0.5 +/- 1.0 diopter (D) (range -3 to 2.4) to 0.06 +/- 0.3 (range -0.9 to 0.6) (P < 0.002) at 6 months. In group 2, the mean preretreatment spherical equivalent was reduced from -0.9 +/- 1.24D (range -3.1 to -0.5) to 0.04 +/- 0.5 (range -1.0 to 1.1) (P < 0.049) at 6 months. At 6 months, UCVA was 20/20 or better in 92% in group 1 and 64% in group 2, of patients, respectively. No eyes lost more than one line of BCVA in group 1 and one eye (7%) lost two lines of BCVA in group 2., Conclusion: Wavefront-guided LASIK retreatment with IR after LASIK is an effective, predictable, and safe procedure in cases requiring a single retreatment. In contrast, eyes with previous retreatments showed less predictability and lower percentage of eyes with postoperative 20/20 UCVA.

Purpose: To report the clinical and histopathologic findings of 2 patients who developed epithelial downgrowth after Descemet stripping automated endothelial keratoplasty (DSAEK)., Methods: A 64-year-old woman (case 1) underwent DSAEK for corneal edema secondary to Fuchs endothelial dystrophy in left eye. However, the graft failed to attach, and a repeat DSAEK was performed 3 weeks later. After 4 months, the patient developed herpes simplex virus keratitis that resulted in anterior stromal scarring. A penetrating keratoplasty was performed 15 months after the initial DSAEK. Our second patient (case 2) was an 87-year-old female who underwent DSAEK for corneal edema secondary to Fuchs endothelial dystrophy in left eye. Six months later, she had an episode of graft rejection and developed secondary glaucoma. At 14 months postoperatively, a retrocorneal membrane was seen involving the temporal half of the endothelial surface of the graft. The retrocorneal membrane extended from the inferior thickened edge of the endothelial keratoplasty graft to the iris stromal surface. An Ahmed shunt implantation followed by repeat DSAEK were then performed. The excised corneal buttons were examined., Results: Histopathologic evaluation showed multilayered epithelium on the interface and attenuated endothelium in the endothelial graft in case 1. The host cornea showed diffuse stromal scarring. Case 2 showed multilayered epithelium with early cyst formation at the edge of the graft. The epithelium extended to involve the endothelial surface without involvement of interface surface. Significant scar formation was observed between the edge of the endothelial keratoplasty graft and thickened host Descemet membrane. Some pigmented cells were present within the epithelial downgrowth. The epithelium stained positively with cytokeratin A1/A3 in both cases., Conclusions: Although rare, epithelial downgrowth can occur after DSAEK and can be associated with graft failure. Early recognition and surgical treatment of epithelial downgrowth is crucial in treating the complications of corneal decompensation and glaucoma.

Purpose: To evaluate postoperative corneal deturgescence after Descemet stripping endothelial keratoplasty (DSAEK) using the Visante anterior segment optical coherence tomography (OCT) system (Carl Zeiss Meditec Inc, Dublin, California, USA)., Design: Retrospective case series., Methods: We included 21 eyes (9 males and 12 females; mean age +/- standard deviation, 76.3 +/- 12 years). The Visante OCT system was used to determine the central and peripheral endothelial keratoplasty graft thickness and total central and peripheral corneal thickness., Results: Central graft thickness decreased from the first day (243.3 +/- 92 microm) to the last visit (147.8 +/- 44 microm; P = .0001). The rate of central graft thinning slowed during the following intervals: during the first week (47 microm), at 1 week to 1 month (40 microm), and at 1 to 6 months (25 microm), with a mild increase at 6 to 9 months (5 microm). Peripheral graft thickness continued to decrease from postoperative day 1 (318.5 +/- 99 microm) to the last visit (196.7 +/- 50 microm; P = .0001). There was a decrease in total central corneal thickness from day 1 (903.8 +/- 179 microm) to the last visit (671 +/- 93 microm; P = .0001). All patients were imaged with the Visante OCT at the first 4 defined postoperative intervals; however, only 9 eyes were imaged at the last interval of 6 to 9 months., Conclusions: After DSAEK, there is a greater thinning of the central graft compared with the peripheral graft. The central cornea thickness decreases and peripheral corneal thickness increases. Central corneal graft deturgescence stabilizes by 6 months after surgery.

Objectives: To report the efficacy and safety profile of sutureless and adhesiveless amniotic membrane device (ProKera, Bio-Tissue, Inc., Miami, FL) in the management of various ocular surface and orbital disorders., Methods: Twenty eyes of 20 patients underwent placement of 21 ProKera implants between March 2006 and July 2007 at the University of Texas Southwestern Medical Center. Patient demographics, indications for placement, and duration of ProKera retention were recorded. Outcomes measured included corneal epithelial healing, visual acuity, patient tolerance, and adverse events., Results: ProKera was most commonly used in patients with corneal neovascularization with or without limbal stem-cell deficiency (10 eyes). Other indications included recurrent pterygium or pseudopterygium (three eyes), anophthalmic orbit contraction (two eyes), persistent epithelial defects (two eyes), severe thinning in a corneal ulcer (one eye), benign hereditary intraepithelial dyskeratosis (one eye), and band keratopathy (one eye). The mean duration of ProKera retention was 25.3 days (range, 0-125) visual acuity improved in 12 eyes (60%). Immediate adverse events included residual epithelial defects after removal (five eyes) and spontaneous extrusion of the implant (four eyes). Six patients (30%) reported eye pain or headache and four eyes (20%) had recurrence of the primary pathology., Conclusions: Sutureless and adhesiveless amniotic membrane transplantation is a safe and effective method to promote healing and reconstruction of the ocular surface and orbit with minimal side effects. Recurrence of the underlying primary pathology remains a concern. The advent of a newer, softer conformer ring may improve patient tolerability and limit discomfort.