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Purpose: To validate a French version of the Visual Function and Corneal Health Status (V-FUCHS) instrument in Fuchs endothelial corneal dystrophy (FECD) and to correlate questionnaire scores with objective measurements of best-corrected visual acuity (BCVA) and intraocular straylight., Methods: The original V-FUCHS 15-item instrument was designed to assess VA and glare in patients with FECD. It was translated from English to French and back translated to English by independent translators. The questionnaire was administered on 2 occasions (4 weeks apart), and BCVA, modified Krachmer grade, and straylight were measured. Construct validity, internal consistency, test-retest reliability, and predictive validity were assessed., Results: A total of 159 subjects, aged 41 to 86 years, were prospectively recruited from July 2019 to October 2020 and classified into 6 groups: mild, moderate, or advanced FECD; unilateral or bilateral Descemet membrane endothelial keratoplasty for FECD; and healthy controls (n = 35). Construct validity was confirmed by item response theory model. Cronbach alpha demonstrated excellent internal consistency (VA: 0.91, glare: 0.88), and intraclass correlations showed good test-retest reliability (VA: 0.75 and glare: 0.82). Significant correlations were observed between both VA and glare factors and the modified Krachmer grade, BCVA, and straylight., Conclusions: The proposed French version of the V-FUCHS instrument provides a valid and reliable tool for the assessment of visual disability in patients with FECD. This questionnaire bridges the gap between self-reported FECD symptoms and objective clinical measurements, which may prove useful to optimize the timing of surgery and to assess postoperative improvement in symptoms., Competing Interests: The authors have no funding or conflicts of interest to disclose., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Purpose: To determine the outcomes and predisposing factors of Descemet's membrane endothelial keratoplasty ( DMEK ) complicated by intraoperative fibrinous reaction., Methods: Retrospective cohort study of 346 DMEKs. Medical charts were reviewed for recipient demographics, surgical indications, donor characteristics, and potential predisposing ocular and systemic factors. For DMEKs complicated by fibrin, surgeons' notes on events leading to fibrin formation and on its intraoperative management, occurrence of graft detachment, primary failure, re-bubbling or regrafting, time to graft clearing, and endothelial cell density were additionally collected., Results: Fifteen (4.3%) DMEKs were complicated by fibrin, which interfered with and protracted graft unfolding in all cases. Median surgical time was longer than for uncomplicated DMEKs ( p  = 0.001). Graft positioning at the end of surgery was suboptimal in seven eyes (47%) and failed in three (20%). Re-bubbling, primary failure, and regraft rates were of 40%, 33% and 53%, respectively. The corneas that cleared did so in three to eight weeks, with median endothelial cell loss of 53% at 12 months. Use of anticoagulants was a preoperative risk factor ( p  = 0.01). Surgeon-identified intraoperative factors included beginner surgeons (87%), prolonged AC shallowing (47%) and graft manipulations (33%), intraocular bleeding (27%), new injector (20%), tight donor scroll (13%), and floppy iris (13%)., Conclusion: Fibrinous reaction is a rare intraoperative complication of DMEK that interferes with graft unfolding and results in poor outcomes. Anticoagulant use appears to be a risk factor and may be compounded by surgical trauma to vascular tissues and prolonged surgical maneuvers.

Purpose: Fuchs endothelial corneal dystrophy (FECD) is characterized by Descemet's membrane (DM) abnormalities, namely an increased thickness and a progressive appearance of guttae and fibrillar membranes. The goal of this study was to identify abnormal extracellular matrix (ECM) proteins expressed in FECD DMs and to evaluate their impact on cell adhesion and migration., Methods: Gene expression profiles from in vitro (GSE112039) and ex vivo (GSE74123) healthy and FECD corneal endothelial cells were analyzed to identify deregulated matrisome genes. Healthy and end-stage FECD DMs were fixed and analyzed for guttae size and height. Immunostaining of fibronectin, tenascin-C, osteopontin, and type XIV collagen was performed on ex vivo specimens, as well as on tissue-engineered corneal endothelium reconstructed using healthy and FECD cells. An analysis of ECM protein expression according to guttae and fibrillar membrane was performed using immunofluorescent staining and phase contrast microscopy. Finally, cell adhesion was evaluated on fibronectin, tenascin-C, and osteopontin, and cell migration was studied on fibronectin and tenascin-C., Results: SPP1 (osteopontin), FN1 (fibronectin), and TNC (tenascin-C) genes were upregulated in FECD ex vivo cells, and SSP1 was upregulated in both in vitro and ex vivo FECD conditions. Osteopontin, fibronectin, tenascin-C, and type XIV collagen were expressed in FECD specimens, with differences in their location. Corneal endothelial cell adhesion was not significantly affected by fibronectin or tenascin-C but was decreased by osteopontin. The combination of fibronectin and tenascin-C significantly increased cell migration., Conclusions: This study highlights new abnormal ECM components in FECD, suggests a certain chronology in their deposition, and demonstrates their impact on cell behavior.

This study aims to examine geometric models of the corneal surface that can be used to reduce in reasonable time the dimensionality of datasets of normal anterior corneas. Polynomial models (P) like Zernike polynomials (ZP) and spherical harmonic polynomials (SHP) were obvious candidates along with their rational function (R) counterparts, namely Zernike rational functions (ZR) and spherical harmonic rational functions (SHR, new model). Knowing that both SHP and ZR were more accurate than ZP for the modeling of normal and keratoconus corneas, it was expected that both spherical harmonic (SH) models (SHP and SHR) would be more accurate than their Zernike (Z) counterparts (ZP and ZR, respectively), and both rational (R) models (SHR and ZR) more accurate than their polynomial counterparts (SHP and ZP, respectively) for a low dimensional space (coefficient number J < 30). This was the case. The SH factor contributed more to accuracy than the R factor. Considering the corneal processing time as a function of J, P models were processed in quasi-linear time with a quasi-null slope and rational models in polynomial time. Z models were faster than SH models, and increasingly so in their R version. In sum, for corneal dimensionality reduction, SHR is the most accurate model, but its processing time is increasingly prohibitive unless the best coefficient combination is identified beforehand. ZP is the fastest model and is reasonably accurate with normal corneas for exploratory tasks. SHP is the best compromise between accuracy and speed., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Purpose: Fuchs endothelial corneal dystrophy (FECD) is characterized by an accelerated depletion of corneal endothelial cells. There is growing evidence that mitochondrial exhaustion is central in the pathology. Indeed, endothelial cells loss in FECD forces the remaining cells to increase their mitochondrial activity, leading to mitochondrial exhaustion. This generates oxidation, mitochondrial damage, and apoptosis, fueling a vicious cycle of cells' depletion. This depletion ultimately causes corneal edema and irreversible loss of transparency and vision. Concurrently to endothelial cells loss, the formation of extracellular mass called guttae on the Descemet's membrane, is a hallmark of FECD. The pathology origins at the center of the cornea and progress outward, like the appearance of guttae., Methods: Using corneal endothelial explants from patients with late-stage FECD at the time of their corneal transplantation, we correlated mitochondrial markers (mitochondrial mass, potential, and calcium) and the level of oxidative stress and apoptotic cells, with the area taken by guttae. The different markers have been analyzed using fluorescent-specific probes and microscopic analysis., Results: We observed a positive correlation between the presence of guttae and the level of mitochondrial calcium and apoptotic cells. We found a negative correlation between the presence of guttae and the level of mitochondrial mass, membrane potential, and oxidative stress., Conclusions: Taken together, these results show that the presence of guttae is correlated with negative outcome in the mitochondrial health, oxidative status, and survival of nearby endothelial cells. This study provides insight on FECD etiology that could lead to treatment targeting mitochondrial stress and guttae.

Fuchs endothelial corneal dystrophy (FECD) is characterized by an accelerated loss of corneal endothelial cells. Since the function of these cells is to maintain the cornea in a state of deturgescence necessary for its transparency, the depletion of corneal endothelial cells ultimately causes corneal edema and irreversible loss of vision. Evidence is accumulating regarding the central involvement of mitochondria in FECD. As we have previously shown, when endothelial cells die and are not replaced, the mitochondria of surviving cells must provide more energy to compensate, leading to a phenomenon we have called mitochondrial burnout. This burnout causes cell death, thus exacerbating an irreversible vicious circle responsible for FECD progression. Corneal transplantation, for which the transplant supply is insufficient, is the only curative alternative for FECD. It thus becomes imperative to find other avenues of treatment. In this article, we tested whether incorporating healthy mitochondria into FECD cells would improve pathological molecular markers of the disease. Using corneal endothelium explants from FECD patients, we demonstrated that incorporation of exogenous mitochondria into FECD cells by co-incubation reduces oxidative stress, increases mitochondrial membrane potential, and reduces mitophagy. In addition, internalization of exogenous mitochondria significantly reduces apoptosis (57% in FECD vs 12% in FECD with internalized mitochondria). Taken together, these results suggest that the internalization of exogenous mitochondria reverses the vicious circle involved in FECD, thus revealing a much-needed novel treatment alternative for FECD., (© 2023. The Author(s).)

Purpose: To evaluate long-term in vivo functionality of corneas regenerated using a cell-free, liquid hydrogel filler (LiQD Cornea) after deep corneal trauma in the feline model. Methods: Two healthy cats underwent 4 mm diameter stepwise 250/450 µm deep surgical corneal ablation with and without needle perforation. The filler comprising 10% (w/w) collagen-like peptide conjugated to polyethylene glycol (CLP-PEG) and 1% fibrinogen and crosslinked with 2% (w/w) 4-(4,6-dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride (DMTMM), was applied to the wound bed previously coated with thrombin (250 U/ml). In situ gelation occurred within 5 min, and a temporary tarsorrhaphy was performed. Eyes were examined weekly for 1 month, then monthly over 12 months. Outcome parameters included slit-lamp, Scheimpflug tomography, optical coherence tomography, confocal and specular microscopy, and immunohistochemistry studies. Results: The gelled filler was seamlessly incorporated, supporting smooth corneal re-epithelialization. Progressive in-growth of keratocytes and nerves into the filler corresponding to the mild haze observed faded with time. The regenerated neo-cornea remained stably integrated throughout the 12 months, without swelling, inflammation, infection, neovascularization, or rejection. The surrounding host stroma and endothelium remained normal at all times. Tomography confirmed restoration of a smooth surface curvature. Conclusion: Biointegration of this hydrogel filler allowed stable restoration of corneal shape and transparency in the feline model, with less inflammation and no neovascularization compared to previous reports in the minipig and rabbit models. It offers a promising alternative to cyanoacrylate glue and corneal transplantation for ulcerated and traumatized corneas in human patients., Competing Interests: MG is a named inventor on PCT/IB 2017/056342 Collagen and CLP-based hydrogels, corneal implants, filler glue and uses thereof, which was assigned to Hyderabad Eye Research Foundation (HERF) and subsequently assigned to North Grove Investments, Inc. by HERF. PCT national phase applications have been filed in the United States, EU, India, China, and Canada. Other authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Juarez, Djallali, Piché, Thériault, Groleau, Beroual, McTiernan, Lin, Hélie, Carrier, Griffith and Brunette.)

SLC4A11 is a H + /NH 3 /water transport protein, of corneal endothelial cells. SLC4A11 mutations cause congenital hereditary endothelial dystrophy and some cases of Fuchs endothelial corneal dystrophy. To probe SLC4A11's roles, we compared gene expression in RNA from corneas of 17-week-old slc4a11 -/- (n = 3) and slc4a11 +/+ mice (n = 3) and subjected to RNA sequencing. mRNA levels for a subset of genes were also assessed by quantitative real-time reverse transcription PCR (qRT RT-PCR). Cornea expressed 13,173 genes, which were rank-ordered for their abundance. In slc4a11 -/- corneas, 100 genes had significantly altered expression. Abundant slc14a1 expression, encoding the urea transporter UT-A, suggests a significant role in the cornea. The set of genes with altered expression was subjected to Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analyses, revealing that alterations clustered into extracellular region, cytoskeleton, cell adhesion and plasma membrane functions. Gene expression changes further clustered into classes (with decreasing numbers of genes): cell fate and development, extracellular matrix and cell adhesion, cytoskeleton, ion homeostasis and energy metabolism. Together these gene changes confirm earlier suggestions of a role of SLC4A11 in ion homeostasis, energy metabolism, cell adhesion, and reveal an unrecognized SLC4A11 role in cytoskeletal organization., (© 2021. The Author(s).)

The long-term survival of biomaterial implants is often hampered by surgery-induced inflammation that can lead to graft failure. Considering that most corneas receiving grafts are either pathological or inflamed before implantation, the risk of rejection is heightened. Here, we show that bioengineered, fully synthetic, and robust corneal implants can be manufactured from a collagen analog (collagen-like peptide-polyethylene glycol hybrid, CLP-PEG) and inflammation-suppressing polymeric 2-methacryloyloxyethyl phosphorylcholine (MPC) when stabilized with the triazine-based crosslinker 4-(4,6-Dimethoxy-1,3,5-triazin-2-yl)-4-methylmorpholinium chloride. The resulting CLP-PEG-MPC implants led to reduced corneal swelling, haze, and neovascularization in comparison to CLP-PEG only implants when grafted into a mini-pig cornea alkali burn model of inflammation over 12 months. Implants incorporating MPC allowed for faster nerve regeneration and recovery of corneal sensation. CLP-PEG-MPC implants appear to be at a more advanced stage of regeneration than the CLP-PEG only implants, as evidenced by the presence of higher amounts of cornea-specific type V collagen, and a corresponding decrease in the presence of extracellular vesicles and exosomes in the corneal stroma, in keeping with the amounts present in healthy, unoperated corneas.

Fuchs endothelial corneal dystrophy (FECD) is characterized by a progressive loss of corneal endothelial cells (CECs) and an abnormal accumulation of extracellular matrix in Descemet's membrane leading to increased thickness and formation of excrescences called guttae. Extracellular matrix homeostasis is modulated by an equilibrium between matrix metalloproteinases (MMPs) and their endogenous tissue inhibitors (TIMPs). This study aimed to investigate MMPs and TIMPs profile in FECD, taking into account cell morphology. Populations of FECD and healthy CECs were cultured and their conditioned media collected for analysis. The presence of proteases in the conditioned media was studied using a semi-quantitative proteome profiler array, and MMPs levels were assessed using quantitative assays (ELISA and quantitative antibody array). MMP activity was determined by zymography and fluorometry. The expression pattern of the membrane type 1-MMP (MT1-MMP, also known as MMP-14) was examined by immunofluorescence on ex vivo FECD and healthy explants of CECs attached to Descemet's membrane. Finally, MMPs and TIMPs protein expression was compared to gene expression obtained from previously collected data. FECD and healthy CEC populations generated cultures of endothelial, intermediate, and fibroblastic-like morphology. Various MMPs (MMP-1, -2, -3, -7, -8, -9, -10, and -12) and TIMPs (TIMP-1 to -4) were detected in both FECD and healthy CECs culture supernatants. Quantitative assays revealed a decrease in MMP-2 and MMP-10 among FECD samples. Both these MMPs can degrade the main extracellular matrix components forming guttae (fibronectin, laminin, collagen IV). Moreover, MMPs/TIMPs ratio was also decreased among FECD cell populations. Activity assays showed greater MMPs/Pro-MMPs proportions for MMP-2 and MMP-10 in FECD cell populations, although overall activities were similar. Moreover, the analysis according to cell morphology revealed among healthy CECs, both increased (MMP-3 and -13) and decreased (MMP-1, -9, -10, and -12) MMPs proteins along with increased MMPs activity (MMP-2, -3, -9, and -10) in the fibroblastic-like subgroup when compared to the endothelial subgroup. However, FECD CECs did not show similar behaviors between the different morphology subgroups. Immunostaining of MT1-MMP on ex vivo FECD and healthy explants revealed a redistribution of MT1-MMP around guttae in FECD explants. At the transcriptional level, no statistically significant differences were detected, but cultured FECD cells had a 12.2-fold increase in MMP1 and a 4.7-fold increase in TIMP3. These results collectively indicate different, and perhaps pathological, MMPs and TIMPs profile in FECD CECs compared to healthy CECs. This is an important finding suggesting the implication of MMPs and TIMPs in FECD pathophysiology., (Copyright © 2021. Published by Elsevier Ltd.)

Purpose: To compare the sealability of femtosecond laser (FSL)-assisted corneal incisions (CIs) with that of triplanar manual (M)-CIs and to determine FSL wound parameters minimizing leakage., Setting: Private practice., Design: Phase IV, single-surgeon, retrospective cohort study., Methods: One eye per patient was included. Two groups defined by the main wound (FSL-CI or M-CI) were compared for leakage, inferred by placement of a suture at the end of surgery. Leakage in FSL-CIs was analyzed as a function of customizable wound parameters: anterior plane depth (APD), posterior plane depth (PPD), anterior side-cut angle (ASCA), and posterior side-cut angle (PSCA). The risk of leakage of FSL-CIs with optimal and nonoptimal parameters was further compared with that of M-CIs., Results: A total of 1100 eyes (757 [68.8%] FSL-CI; 343 [31.2%] M-CI) were included. Wound leakage occurred in 133 FSL-CI (17.6%) and 30 M-CI eyes (8.7%) (P < .001). FSL wound parameters associated with the lowest risk of leakage were 60% APD, 70% PPD, 120 degrees ASCA, and 70 degrees PSCA. FSL-CIs constructed with at least 3 optimal parameters (60% APD, 70% PPD, and 120 degrees ASCA) had a similar risk of leakage to M-CIs (odds ratio [OR], 1.1; 95% CI, 0.5-2.3). FSL-CIs with suboptimal parameters had twice the risk of leakage of M-CIs (OR, 2.0; 95% CI, 1.1-3.8)., Conclusions: Overall, FSL-CIs leaked more than M-CIs. However, FSL-CIs with optimized wound profiles had an equivalent risk of leakage to M-CIs. Wound parameter customization is an asset of FSL technology that allows optimization of FSL-CI sealability.

Fuchs endothelial corneal dystrophy (FECD) is a degenerative eye disease characterized by corneal endothelial cell (CEC) death and the formation of guttae, an abnormal thickening of CEC's basement membrane. At the tissue level, an oxidative stress causing mitochondrial damage and CEC death have been described to explain FECD pathogenesis. At the cellular level, our group has previously observed significant variability in the mitochondrial mass of FECD CECs. This led us to hypothesize that mitochondrial mass variability might play a key role in the chronology of events eventually leading to CEC death in FECD. We thus used different fluorescent markers to assess mitochondrial health and functionality as a function of mitochondrial mass in FECD corneal endothelial explants, namely, intra-mitochondrial calcium, mitochondrial membrane potential, oxidation level and apoptosis. This has led us to describe for the first time a sequence of events leading to what we referred to as a mitochondrial burnout, and which goes as follow. FECD CECs initially compensate for endothelial cell losses by incorporating mitochondrial calcium to help generating more ATP, but this leads to increased oxidation. CECs then resist the sustained need for more ATP by increasing their mitochondrial mass, mitochondrial calcium and mitochondrial membrane potential. At this stage, CECs reach their maximum capacity and start to cope with irreversible oxidative damage, which leads to mitochondrial burnout. This burnout is accompanied by a dissipation of the membrane potential and a release of mitochondrial calcium, which in turn leads to cell death by apoptosis.

Purpose: To report outcomes of keratopigmentation (KP) with commercial black ink in the treatment of dysphotopsia secondary to laser peripheral iridotomies (LPI) using manual anterior stromal puncture (ASP) and manual lamellar pocket (LP)., Methods: This is a retrospective case series of eyes that underwent KP for treatment of dysphotopsia secondary to LPI. Patients' postoperative symptoms were categorized as resolved, improved, no change, or worse. Any intraoperative and postoperative complications were noted, as well as the need for further treatments., Results: Five eyes in 4 patients underwent ASP, and 14 eyes in 13 patients underwent LP. Only 1 patient had improvement in symptoms in the AK group. Four patients had a complete resolution of symptoms after LP, whereas 7 had symptomatic improvement and 3 did not notice any change. Fifty-five percent of patients in the LP group experienced late-onset depigmentation between 3 and 5 years postoperatively. One patient who underwent 2 LP re-treatments experienced irregular corneal steepening with nonprogressive corneal thinning. Overall, there were no serious adverse reactions to the pigment used., Conclusions: ASP was not suitable for the treatment of dysphotopsia secondary to LPI. In the short term, LP had good outcomes, but commercial black ink was prone to depigmentation at 3 to 5 years postoperatively. We therefore do not recommend the use of such pigment for long-term management of dysphotopsia secondary to LPI.

This is a case report of post-laser in situ keratomileusis (LASIK) multidrug-resistant Mycobacterium abscessus keratitis managed with combined topical amikacin and linezolid, flap amputation, and corticosteroids. A 34-year-old woman presented with a corneal interface infiltrate 3 weeks after LASIK. Cultures isolated mycobacteria. The infiltrate did not improve under intensive topical therapy and interface irrigation with empiric antibiotics over 5 weeks, and the infiltrate progressed to severe inflammation and stromal neovascularization. After identification of M abscessus susceptible only to amikacin and linezolid, antimicrobials were adjusted and the flap was ablated. Cultures repeated 1 week later came back negative. However, stromal inflammation and neovascularization persisted. Topical steroids achieved regression of the inflammation within 1 week. Identification of the mycobacterial pathogen and its susceptibilities is essential given the possibility of multidrug resistance. Topical linezolid can be effective in susceptible species. Corticosteroids can be helpful in cases with severe inflammation., (Copyright © 2019. Published by Elsevier Inc.)

Cells and tissues are influenced by environmental conditions. In vivo, the corneal endothelium is subjected to hydrostatic intraocular pressure (IOP) and to the hydrokinetic pressure of the moving aqueous humor in the anterior chamber. In this paper, we used a corneal bioreactor to recreate the IOP condition and investigated the effect of the in vivo hydrodynamic environment of corneal endothelial cells on the formation of tight junctions. Native ex vivo corneas and engineered corneal endothelia subjected to pressure showed an increase in ZO-1 expression at the cell periphery. Pressure also improved the corneal transparency of engineered and native corneas. Corneal thickness was accordingly reduced from 926 ± 70 μm to 651 ± 70 μm for the engineered corneal endothelium and from 847 ± 27 μm to 571 ± 23 μm for the native endothelium. These results suggest that the hydrodynamic pressure of the anterior chamber is important for the cell junction integrity of the corneal endothelium., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

This study investigates the possible existence of different natural corneal shape categories. This is important to better describe cornea for both diagnostic and therapeutic assessments. We started by describing corneal shape of different populations as a function of influencing clinical data i.e. age, ametropia and gender. This was done by averaging Zernike polynomial (ZP) decomposition of the anterior surfaces in each subgroup. The results showed small but significant differences of shape that are supported by the literature. This motivated us to examine the feasibility of characterizing the normal corneal shape with an automatic method of clustering independent of any clinical a priori knowledge. Since we did not know beforehand the number of corneal categories, agglomerative hierarchical clustering was applied on ZP coefficients for a large database. The dendrogram based on the Ward's distance was evaluated with two different clustering validity indexes (coefficient of determination R 2 and semi partial R 2 (SPR 2 ). The optimal number of categories was around four showing corneal shapes ranging from flatter to steeper.

Fuchs endothelial corneal dystrophy (FECD) is a corneal pathology that affects the endothelial cell's ability to maintain deturgescence, resulting in a progressive loss of corneal transparency. In this study, we investigated the expression of function-related proteins in corneal endothelial cells using FECD or healthy corneal endothelial cells, either in a cell culture two-dimensional model or in an engineered corneal endothelium three-dimensional tissue model. No statistically significant difference in gene regulation was observed for the function-related families ATP1, SLC4, SLC16, AQP, TJP, and CDH between the FECD and the healthy cell models. Similarly, no difference in barrier integrity (transendothelial electrical resistance measurements and permeability assays) was observed in vitro between FECD and healthy cultured cells. Protein expression of the key function-related families was decreased for Na + /K + -ATPase α1 subunit, monocarboxylate transporters 1 and 4 in native ex vivo end-stage FECD specimens, whereas it returned to levels comparable to that of healthy tissues in the engineered FECD model. These results indicate that cell expansion and tissue engineering culture conditions can generate a corneal endothelium from pathologic FECD cells, with levels of function-related proteins similar to that of healthy tissues. Overall, these results explain why it is possible to reform a functional endothelium using corneal endothelial cells isolated from nonfunctional FECD pathologic specimens., (Copyright © 2018 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

Primary corneal endothelial cell (CEC) cultures and 3D-engineered tissue models were used to study the aberrant deposition of extracellular matrix (ECM) in a vision impairing pathology known as Fuchs endothelial corneal dystrophy (FECD). CECs were isolated from excised Descemet membranes of patients with end-stage FECD. CECs isolated from healthy corneas served as controls. Microarray gene profiling was performed on postconfluent cultures of healthy and FECD cells. Protein expression analyses were conducted on tissue models that were engineered by seeding an endothelium on previously devitalized human stromal carriers. The engineered endothelia were kept in culture for 1-3 weeks to reform the endothelial monolayer. Protein expression of integrin subunits α4, α6, αv, and β1, as well as laminin, type IV collagen, fibronectin, clusterin, and transforming growth factor β-induced protein (TGFβIp) was then assessed by immunofluorescence. Microarray analysis showed nonstatistical twofold downregulation of collagen-coding genes (COL4A4, COL8A2, and COL21A1) and a twofold upregulation of the COL6A1, laminin α3 gene LAMA3, and integrin subunit α10 gene ITGA10 in FECD cells. Fibronectin type III domain containing 4 (FNDC4) and integrin β5 (ITGB5) genes was significantly upregulated in FECD cells. Immunostainings demonstrated that the protein expression of the integrin subunits α4, α6, αv, and β1, type IV collagen, as well as laminin remained similar between native and engineered endothelia. TGFβIp expression was found on the stromal side of both FECD and healthy Descemet's membrane, and only one out of three FECD specimens was positive for the clusterin protein. Interestingly, the ECM protein fibronectin was also found to have a stronger presence on engineered FECD tissues, a result consistent with the native FECD specimens. To conclude, this study allowed to identify fibronectin deposition as one of the first steps in the pathogenesis of FECD, as defined by our engineered tissue model. This opens the way to an entirely new perspective for in vitro pharmacological testing of new therapies for FECD, the leading indication for corneal transplantation in North America.

Corneal blindness is a major cause of blindness in the world and corneal transplantation is the only widely accepted treatment to restore sight in these eyes. However, it is becoming increasingly difficult for eye banks to meet the increasing demand for transplantable tissue, which is in part due to population aging. Donor tissue shortage is therefore a growing concern globally and there is a need for alternatives to human donor corneas. Biosynthetic corneal substitutes offer several significant advantages over native corneas: Large-scale production offers a powerful potential solution to the severe shortage of human donor corneas worldwide; Good manufacturing practices ensure sterility and quality control; Acellular corneal substitutes circumvent immune rejection induced by allogeneic cells; Optical and biomechanical properties of the implants can be adapted to the clinical need; and finally these corneal substitutes could benefit from new advances in biomaterials science, such as surface coating, functionalization and nanoparticles. This review highlights critical contributions from laboratories working on corneal stromal substitutes. It focuses on synthetic inert prostheses (keratoprostheses), acellular scaffolds with and without enhancement of endogenous regeneration, and cell-based replacements. Accent is put on the physical properties and biocompatibility of these biomaterials, on the functional and clinical outcome once transplanted in vivo in animal or human eyes, as well as on the main challenges of corneal stromal replacement. Regulatory and economic aspects are also discussed. All of these perspectives combined highlight the founding principles of the clinical application of corneal stromal replacement, a concept that has now become reality., (Copyright © 2017 Elsevier Ltd. All rights reserved.)