rhFGF-21 accelerates corneal epithelial wound healing through the attenuation of oxidative stress and inflammatory mediators in diabetic mice.

Diabetic keratopathy, commonly associated with a hyperactive inflammatory response, is one of the most common eye complications of diabetes. The peptide hormone fibroblast growth factor-21 (FGF-21) has been demonstrated to have anti-inflammatory and antioxidant properties. However, whether administration of recombinant human (rh) FGF-21 can potentially regulate diabetic keratopathy is still unknown. Therefore, in this work, we investigated the role of rhFGF-21 in the modulation of corneal epithelial wound healing, the inflammation response, and oxidative stress using type 1 diabetic mice and high glucose-treated human corneal epithelial cells. Our experimental results indicated that the application of rhFGF-21 contributed to the enhancement of epithelial wound healing. This treatment also led to advancements in tear production and reduction in corneal edema. Moreover, there was a notable reduction in the levels of proinflammatory cytokines such as TNF-α, IL-6, IL-1ß, MCP-1, IFN-γ, MMP-2, and MMP-9 in both diabetic mouse corneal epithelium and human corneal epithelial cells treated with high glucose. Furthermore, we found rhFGF-21 treatment inhibited reactive oxygen species production and increased levels of anti-inflammatory molecules IL-10 and SOD-1, which suggests that FGF-21 has a protective role in diabetic corneal epithelial healing by increasing the antioxidant capacity and reducing the release of inflammatory mediators and matrix metalloproteinases. Therefore, we propose that administration of FGF-21 may represent a potential treatment for diabetic keratopathy.

The role of the JAK/STAT3 signaling pathway in acquired corneal diseases.

Acquired corneal diseases such as dry eye disease (DED), keratitis and corneal alkali burns are significant contributors to vision impairment worldwide, and more effective and innovative therapies are urgently needed. The Janus kinase/signal transducer and activator of transcription 3 (JAK/STAT3) signaling pathway plays an indispensable role in cell metabolism, inflammation and the immune response. Studies have shown that regulators of this pathway are extensively expressed in the cornea, inducing significant activation of JAK/STAT3 signaling in specific acquired corneal diseases. The activation of JAK/STAT3 signaling contributes to various pathophysiological processes in the cornea, including inflammation, neovascularization, fibrosis, and wound healing. In the context of DED, the hypertonic environment activates JAK/STAT3 signaling to stimulate corneal inflammation. Inflammation and injury progression in infectious keratitis can also be modulated by JAK/STAT3 signaling. Furthermore, JAK/STAT3 signaling is involved in every stage of corneal repair after alkali burns, including acute inflammation, angiogenesis and fibrosis. Treatments modulating JAK/STAT3 signaling have shown promising results in attenuating corneal damage, indicating its potential as a novel therapeutic target. Thus, this review emphasizes the multiple roles of the JAK/STAT3 signaling pathway in common acquired corneal disorders and summarizes the current achievements of JAK/STAT3-targeting therapy to provide new insights into future applications.

Effects of hypoxia in the diabetic corneal stroma microenvironment.

Corneal dysfunctions associated with Diabetes Mellitus (DM), termed diabetic keratopathy (DK), can cause impaired vision and/or blindness. Hypoxia affects both Type 1 (T1DM) and Type 2 (T2DM) surprisingly, the role of hypoxia in DK is unexplored. The aim of this study was to examine the impact of hypoxia in vitro on primary human corneal stromal cells derived from Healthy (HCFs), and diabetic (T1DMs and T2DMs) subjects, by exposing them to normoxic (21% O2) or hypoxic (2% O2) conditions through 2D and 3D in vitro models. Our data revealed that hypoxia affected T2DMs by slowing their wound healing capacity, leading to significant alterations in oxidative stress-related markers, mitochondrial health, cellular homeostasis, and endoplasmic reticulum health (ER) along with fibrotic development. In T1DMs, hypoxia significantly modulated markers related to membrane permeabilization, oxidative stress via apoptotic marker (BAX), and protein degradation. Hypoxic environment induced oxidative stress (NOQ1 mediated reduction of superoxide in T1DMs and Nrf2 mediated oxidative stress in T2DMs), modulation in mitochondrial health (Heat shock protein 27 (HSP27), and dysregulation of cellular homeostasis (HSP90) in both T1DMs and T2DMs. This data underscores the significant impact of hypoxia on the diabetic cornea. Further studies are warranted to delineate the complex interactions.

Wnt/ß-catenin signaling in corneal epithelium development, homeostasis, and pathobiology.

The corneal epithelium is located on the most anterior surface of the eyeball and protects against external stimuli. The development of the corneal epithelium and the maintenance of corneal homeostasis are essential for the maintenance of visual acuity. It has been discovered recently via the in-depth investigation of ocular surface illnesses that the Wnt/ß-catenin signaling pathway is necessary for the growth and stratification of corneal epithelial cells as well as the control of endothelial cell stability. In addition, the Wnt/ß-catenin signaling pathway is directly linked to the development of common corneal illnesses such as keratoconus, fungal keratitis, and corneal neovascularization. This review mainly summarizes the role of the Wnt/ß-catenin signaling pathway in the development, homeostasis, and pathobiology of cornea, hoping to provide new insights into the study of corneal epithelium and the treatment of related diseases.

Amelioration of Fibrosis via S1P Inhibition Is Regulated by Inactivation of TGF-ß and SPL Pathways in the Human Cornea.

Human corneal fibrosis can lead to opacity and ultimately partial or complete vision loss. Currently, corneal transplantation is the only treatment for severe corneal fibrosis and comes with the risk of rejection and donor shortages. Sphingolipids (SPLs) are known to modulate fibrosis in various tissues and organs, including the cornea. We previously reported that SPLs are tightly related to both, transforming growth factor beta (TGF-ß) signaling and corneal fibrogenesis. The aim of this study was to investigate the effects of sphingosine-1-phosphate (S1P) and S1P inhibition on specific TGF-ß and SPL family members in corneal fibrosis. Healthy human corneal fibroblasts (HCFs) were isolated and cultured in EMEM + FBS + VitC (construct medium) on 3D transwells for 4 weeks. The following treatments were prepared in a construct medium: 0.1 ng/mL TGF-ß1 (ß1), 1 µM sphingosine-1-phosphate (S1P), and 5 µM Sphingosine kinase inhibitor 2 (I2). Five groups were tested: (1) control (no treatment); rescue groups; (2) ß1/S1P; (3) ß1/I2; prevention groups; (4) S1P/ß1; and (5) I2/ß1. Each treatment was administered for 2 weeks with one treatment and switched to another for 2 weeks. Using Western blot analysis, the 3D constructs were examined for the expression of fibrotic markers, SPL, and TGF-ß signaling pathway members. Scratch assays from 2D cultures were also utilized to evaluate cell migration We observed reduced fibrotic expression and inactivation of latent TGF-ß binding proteins (LTBPs), TGF-ß receptors, Suppressor of Mothers Against Decapentaplegic homologs (SMADs), and SPL signaling following treatment with I2 prevention and rescue compared to S1P prevention and rescue, respectively. Furthermore, we observed increased cell migration following stimulation with I2 prevention and rescue groups, with decreased cell migration following stimulation with S1P prevention and rescue groups after 12 h and 18 h post-scratch. We have demonstrated that I2 treatment reduced fibrosis and modulated the inactivation of LTBPs, TGF-ß receptors, SPLs, and the canonical downstream SMAD pathway. Further investigations are warranted in order to fully uncover the potential of utilizing SphK I2 as a novel therapy for corneal fibrosis.

Exome sequencing and functional studies in zebrafish identify WDR8 as the causative gene for isolated Microspherophakia in Indian families.

Isolated Microspherophakia (MSP) is an autosomal recessive disorder characterized by a smaller than normal spherical lens. Till date, LTBP2 is the only gene shown to cause MSP. We used homozygosity mapping and whole-exome sequencing and identified a homozygous mutation, c.1148C > T (p.Pro383Leu), in the WDR8 (or WRAP73) gene in two Indian MSP families. In vitro experiments showed that the missense mutation renders the protein unstable. WDR8 is a centriolar protein that has important roles in centrosomal assembly, spindle pole formation and ciliogenesis. Co-immunoprecipitation experiments from HeLa cells indicated that the mutation interferes with the interaction of WDR8 with its binding partners. In zebrafish, both morpholino-mediated knockdown and CRISPR/Cas knockout of wdr8 resulted in decreased eye and lens size. The lack of wdr8 affected cell cycle progression in the retinal cells, causing a reduction in cell numbers in the retina and lens. The reduction in eye size and the cell cycle defects were rescued by exogenous expression of the human wild-type WDR8. However, the human mutant WDR8 (p.Pro383Leu) was unable to rescue the eye defects, indicating that the missense mutation abrogates WDR8 protein function. Thus, our zebrafish results suggested that WDR8 is the causative gene for MSP in these Indian families.

Evaluation of the Algerbrush II rotating burr as a tool for inducing ocular surface failure in a mouse model.

Purpose: The Algerbrush II has been widely used to induce corneal and limbal injuries in animal models. The extent of injury varies with the duration of exposure, pressure from the placement of the burr, and the size of the burr. However, no study has explored the correlation between the duration of exposure and the severity of injury in mouse model with corneal and limbal stem cell deficiency (LSCD) induced using the Algerbrush II. Therefore, this study aimed to evaluate the variations in the severity of corneal and limbal injury with different durations of the Algerbrush II application. Methods: The entire cornea and limbus of C57BL/6 mice were injured for 30-45 s, 60-75 s, 90-120 s, and 3-4 min. Photography and slit-lamp examination was performed on days 0, 2, 4, and 7, followed by hematoxylin & eosin, periodic acid-Schiff, and immunohistochemical staining. Statistical analysis was performed using one way ANOVA analysis. Results: A duration of 30-45 s of injury was found to be sufficient to induce superficial corneal and limbal epithelial debridement and re-epithelialization was completed in all eyes by day 7; however, clinical signs of LSCD were not observed in all mice. Increasing the exposure time to 90-120 s resulted in central 2+ corneal opacity with limbal and paracentral corneal neovascularization. All eyes injured for 3-4 min displayed clinical signs of LSCD, such as persistent epithelial defects on day 7 after the injury, central corneal neovascularization, and 2.2+ diffuse corneal opacity. Histological signs of LSCD, including goblet cell metaplasia and K13 expression on the corneal surface, were observed in all injured eyes. Conclusions: Our findings suggest that the duration of injury is an important factor influencing the severity of LSCD in a murine model of injury. A 1-mm rotating burr was found to be more effective for keratectomy and pigment release, whereas a 0.5-mm burr was more suitable for corneal epithelial debridement.

Proteomic characterization of aqueous humor in corneal endothelial decompensation after penetrating keratoplasty.

Corneal endothelial decompensation (CED) is the major cause of the long-term graft failure, but the underlying mechanisms remain unclear. The purpose of this study was to characterize the proteomic profile in CED aqueous humor (AH) after penetrating keratoplasty (PKP). We collected AH samples (n = 6/group) from CED patients underwent PKP and cataract patients, respectively. The label-free quantitative proteomic analysis was performed to identify the differentially-expressed proteins (DEPs). The biological functions of DEPs were evaluated using Gene Ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genome (KEGG) analysis. The protein-protein interaction (PPI) network construction was employed to distinguish the hub proteins of DEPs, and the selected proteins were validated by parallel reaction monitoring (PRM). The human peripheral blood mononuclear cells (PBMCs) were adopted to investigate the effect of biglycan (BGN) on inflammatory response, and the subsequent outcomes of inflammation on human corneal endothelial cells (HCECs). A total of 174 DEPs were identified in CED AH of patients underwent PKP, including 102 up-regulated proteins and 72 down-regulated proteins. Bioinformatics analysis revealed the significant enrichment of cytokine-mediated signaling pathway and extracellular matrix (ECM) organization in the up-regulated proteins, as well as the alterations of cellular components, including the increase of collagen and complement component C1 complex, and reduction in extracellular exosomes. A hub protein cluster of 15 proteins was determined by Molecular Complex Detection (MCODE), including FN1, BGN, COMP, COL11A1, COLA3A1, and COL1A1. Moreover, BGN promoted pro-inflammatory cytokine (such as TNF-α, IL-1ß and IL-6) production in PBMCs through NF-κB signaling pathway, which subsequently resulted in HCECs death. These findings provided a systemic protein profile of AH in CED patients after corneal transplantation, with the alterations implicated in cytokine-mediated signaling, ECM, complement system, and exsomes. The identified proteins and signaling pathways probably paved the novel insight into understanding the pathogenesis of the disease.

FoxC1 activates limbal epithelial stem cells following corneal epithelial debridement.

Limbal epithelial stem cells are not only critical for corneal epithelial homeostasis but also have the capacity to change from a relatively quiescent mitotic phenotype to a rapidly proliferating cell in response to population depletion following corneal epithelial wounding. Pax6+/- mice display many abnormalities including corneal vascularization and these aberrations are consistent with a limbal stem cell deficiency (LSCD) phenotype. FoxC1 has an inhibitory effect on corneal avascularity and a positive role in stem cell maintenance in many tissues. However, the role of FoxC1 in limbal epithelial stem cells remains unknown. To unravel FoxC1's role(s) in limbal epithelial stem cell homeostasis, we utilized an adeno-associated virus (AAV) vector to topically deliver human FOXC1 proteins into Pax6 +/- mouse limbal epithelium. Under unperturbed conditions, overexpression of FOXC1 in the limbal epithelium had little significant change in differentiation (PAI-2, Krt12) and proliferation (BrdU, Ki67). Conversely, such overexpression resulted in a marked increase in the expression of putative limbal epithelial stem cell markers, N-cadherin and Lrig1. After corneal injuries in Pax6 +/- mice, FOXC1 overexpression enhanced the behavior of limbal epithelial stem cells from quiescence to a highly proliferative status. Overall, the treatment of AAV8-FOXC1 may be beneficial to the function of limbal epithelial stem cells in the context of a deficiency of Pax6 function.

Defective perlecan-associated basement membrane regeneration and altered modulation of transforming growth factor beta in corneal fibrosis.

In the cornea, the epithelial basement membrane (EBM) and corneal endothelial Descemet's basement membrane (DBM) critically regulate the localization, availability and, therefore, the functions of transforming growth factor (TGF)ß1, TGFß2, and platelet-derived growth factors (PDGF) that modulate myofibroblast development. Defective regeneration of the EBM, and notably diminished perlecan incorporation, occurs via several mechanisms and results in excessive and prolonged penetration of pro-fibrotic growth factors into the stroma. These growth factors drive mature myofibroblast development from both corneal fibroblasts and bone marrow-derived fibrocytes, and then the persistence of these myofibroblasts and the disordered collagens and other matrix materials they produce to generate stromal scarring fibrosis. Corneal stromal fibrosis often resolves completely if the inciting factor is removed and the BM regenerates. Similar defects in BM regeneration are likely associated with the development of fibrosis in other organs where perlecan has a critical role in the modulation of signaling by TGFß1 and TGFß2. Other BM components, such as collagen type IV and collagen type XIII, are also critical regulators of TGF beta (and other growth factors) in the cornea and other organs. After injury, BM components are dynamically secreted and assembled through the cooperation of neighboring cells-for example, the epithelial cells and keratocytes for the corneal EBM and corneal endothelial cells and keratocytes for the corneal DBM. One of the most critical functions of these reassembled BMs in all organs is to modulate the pro-fibrotic effects of TGFßs, PDGFs and other growth factors between tissues that comprise the organ.

Early Clinical Outcomes of the First Commercialized Human Autologous Ex Vivo Cultivated Oral Mucosal Epithelial Cell Transplantation for Limbal Stem Cell Deficiency: Two Case Reports and Literature Review.

The first product in the world for ex vivo cultivated oral mucosal epithelial cell transplantation (COMET) to treat limbal stem cell deficiency (LSCD), named Ocural®, was launched in June 2021 in Japan. COMET was performed on two patients, including the first case in the post-marketing phase of Ocural®. Pathological and immunohistochemical examinations were also carried out using specimens obtained before and after COMET and the spare cell sheet. In case 1, the ocular surface remained free from epithelial defects for approximately six months. In case 2, although defect of the cornea-like epithelia was observed after COMET for one month, it was resolved after the insertion of lacrimal punctal plugs. In case 1, adjuvant treatment was interrupted due to an accident during the second month after COMET, resulting in conjunctival ingrowth and corneal opacity. Eventually, a lamellar keratoplasty was required at six months after COMET. Immunohistochemistry revealed the presence of markers for stem cells (p63, p75), proliferation (Ki-67), and differentiation (Keratin-3, -4, and -13) in both the cornea-like tissue after COMET and a cultivated oral mucosal epithelial cell sheet. In conclusion, Ocural® can be accomplished without major complications, and the stem cells derived from oral mucosa might be successfully engrafted.

Mesenchymal Stem Cells and Exosomes: A Novel Therapeutic Approach for Corneal Diseases.

The cornea, with its delicate structure, is vulnerable to damage from physical, chemical, and genetic factors. Corneal transplantation, including penetrating and lamellar keratoplasties, can restore the functions of the cornea in cases of severe damage. However, the process of corneal transplantation presents considerable obstacles, including a shortage of available donors, the risk of severe graft rejection, and potentially life-threatening complications. Over the past few decades, mesenchymal stem cell (MSC) therapy has become a novel alternative approach to corneal regeneration. Numerous studies have demonstrated the potential of MSCs to differentiate into different corneal cell types, such as keratocytes, epithelial cells, and endothelial cells. MSCs are considered a suitable candidate for corneal regeneration because of their promising therapeutic perspective and beneficial properties. MSCs compromise unique immunomodulation, anti-angiogenesis, and anti-inflammatory properties and secrete various growth factors, thus promoting corneal reconstruction. These effects in corneal engineering are mediated by MSCs differentiating into different lineages and paracrine action via exosomes. Early studies have proven the roles of MSC-derived exosomes in corneal regeneration by reducing inflammation, inhibiting neovascularization, and angiogenesis, and by promoting cell proliferation. This review highlights the contribution of MSCs and MSC-derived exosomes, their current usage status to overcome corneal disease, and their potential to restore different corneal layers as novel therapeutic agents. It also discusses feasible future possibilities, applications, challenges, and opportunities for future research in this field.

Direct oral mucosal epithelial transplantation supplies stem cells and promotes corneal wound healing to treat refractory persistent corneal epithelial defects.

Persistent corneal epithelial defects (PED) can lead to irreversible blindness, seriously affecting the social function and life quality of these patients. When it comes to refractory PED, such as limbal stem cell deficiency (LSCD), that does not respond to standard managements, stem cell therapy is an ideal method. Oral mucosal epithelium (OME) abundant with stem cells within the base, is a promising autologous biomaterial, with much resemblance to corneal epithelial structures. In this experiment, uncultured autologous rat OME was directly applied to alkali burned corneas. Clinical evaluations and histological analyses showed that the transplantation accelerated the healing process, presenting faster re-epithelization and better formation of corneal epithelial barrier. To further investigate the therapeutic mechanism, oral epithelium was transplanted to de-epithelialized cornea in vitro for organ culture. It could be observed that the oral epithelial cells could migrate to the corneal surface and form smooth and stratified epithelium. Immunofluorescence staining results showed that the re-formed epithelium derived from OME, maintained stemness and transformed to corneal epithelial phenotype to some extent. Corneal stroma may provide the suitable microenvironment to promote the trans-differentiation of oral stem cells. Thus, both in vivo and in vitro experiments suggested that oral epithelium could play a positive role in treating refractory PED.

Topical losartan inhibits corneal scarring fibrosis and collagen type IV deposition after Descemet's membrane-endothelial excision in rabbits.

The purpose of this study was to examine the effect of topical and/or oral angiotensin converting enzyme II inhibitor and TGF-beta signaling blocker losartan on corneal stromal fibrosis that developed in rabbit corneas after Descemetorhexis removal of central Descemet's membrane and corneal endothelium. Twenty-eight New Zealand white rabbits were included and either had 8 mm central Descemetorhexis or sham control surgery without Descemetorhexis in one eye. Groups of 4 eyes without Descemetorhexis were treated for one month with no medications, topical losartan or oral losartan. Groups of 4 eyes with Descemetorhexis were treated with topical and oral vehicle, topical losartan, oral losartan, or both topical losartan and oral losartan for one month. Standardized slit lamp photos were obtained with central opacity intensity measured with ImageJ. The posterior fibrotic zone of corneas was measured on immunohistochemistry for alpha-smooth muscle actin (SMA) and keratocan using QuPath analysis. Collagen type IV expression in the posterior cornea was quantitated with ImageJ and duplex immunohistochemistry for collagen type IV and TGF beta-1. After Descemetorhexis, topical, but not oral, losartan decreased the intensity of central stromal opacity, reduced peripheral corneal scarring, and decreased alpha-smooth muscle actin myofibroblast fibrosis area compared to corneas that had Descemetorhexis and treatment with vehicles alone. Topical losartan decreased posterior stromal cellular, non-Descemet's membrane, collagen type IV production, that is likely stimulated by TGF beta as part of a negative regulatory feedback mechanism, compared to vehicle treatment at one month after Descemetorhexis. Topical losartan is likely to be effective in reducing corneal scarring fibrosis produced by traumatic injury, microbial infection, and some corneal diseases and surgeries.

The Role of Ectodysplasin A on the Ocular Surface Homeostasis.

Ectodysplasin A (EDA), a ligand of the TNF family, plays an important role in maintaining the homeostasis of the ocular surface. EDA is necessary for the development of the meibomian gland, the lacrimal gland, as well as the proliferation and barrier function of the corneal epithelium. The mutation of EDA can induce the destruction of the ocular surface resulting in keratopathy, abnormality of the meibomian gland and maturation of the lacrimal gland. Experimental animal studies showed that a prenatal ultrasound-guided intra-amniotic injection or postnatal intravenous administration of soluble recombinant EDA protein can efficiently prevent the development of ocular surface abnormalities in EDA mutant animals. Furthermore, local application of EDA could restore the damaged ocular surface to some extent. Hence, a recombinant EDA-based therapy may serve as a novel paradigm to treat ocular surface disorders, such as meibomian gland dysfunction and corneal epithelium abnormalities.

SOX2 Is a Univocal Marker for Human Oral Mucosa Epithelium Useful in Post-COMET Patient Characterization.

Total bilateral Limbal Stem Cells Deficiency is a pathologic condition of the ocular surface due to loss or impairment of corneal stem cell function, altering homeostasis of the corneal epithelium. Cultivated Oral Mucosa Epithelial Transplantation (COMET) is the only autologous treatment for this pathology. During the follow-up, a proper characterization of the transplanted oral mucosa on the ocular surface supports understanding the regenerative process. The previously proposed markers for oral mucosa identification (e.g., keratins 3 and 13) are co-expressed by corneal and conjunctival epithelia. Here, we propose a new specific marker to distinguish human oral mucosa from the epithelia of the ocular surface. We compared the transcriptome of holoclones (stem cells) from the human oral mucosa, limbal and conjunctival cultures by microarray assay. High expression of SOX2 identified the oral mucosa vs. cornea and conjunctiva, while PAX6 was highly expressed in corneal and conjunctival epithelia. The transcripts were validated by qPCR, and immunological methods identified the related proteins. Finally, the proposed markers were used to analyze a 10-year follow-up aniridic patient treated by COMET. These findings will support the follow-up analysis of COMET treated patients and help to shed light on the mechanism of corneal repair and regeneration.

Aldehyde Dehydrogenases Expression in Corneal Epithelial Cells with Limbal Stem Cell Deficiency.

PURPOSE: The purpose of the present study is to investigate the expression of aldehyde dehydrogenases (ALDHs) in rabbit corneas with limbal stem cell deficiency (LSCD) and corneas treated with cultured autologous oral mucosa epithelial cell sheet CAOMECS designed to reconstruct the ocular surface with LSCD. METHODS: New Zealand white rabbit autologous oral mucosal epithelial cells were isolated from a buccal biopsy and cultured to be grafted back onto corneas of rabbit model of LSCD. Immunofluorescent staining and Western blot analysis were used to compare the expression of ALDH1A1 and ALDH1A3 in healthy, LSCD-diseased, CAOMECS treated corneas. Human oral mucosal and corneal epithelial cells (OMECS and CECs) were cultured and treated with retinoic acid (RA) to further investigate the expression of ALDHs. RESULTS: In healthy corneas, ALDH1A1 and ALDH1A3 were markedly expressed in basal cells of corneal epithelium. In LSCD diseased corneas, ALDH1A1 and ALDH1A3 were markedly expressed in the conjunctivalized apical epithelial cells, the goblet cells, and the stroma. CAOMECS grafted corneas showed a decreased expression of ALDHs as compared to LSCD diseased corneas. Western blot analysis confirmed the up regulation of ALDH1A1 and ALDH1A3 expression in LSCD-diseased corneal epithelial cells. CAOMECS expressed low levels of ALDH1A1 and ALDH1A3, as compared to diseased CECs (D-CEC). When ALDH1A3 was up regulated by retinoic acid treatment in OMECS, Pax-6 expression was down regulated, suggesting a decrease in regenerative capacity when ALDH enzymes are up regulated. CONCLUSIONS: These findings report for the first time the up regulation of ALDH1A1 and ALDH1A3 in rabbit corneas with LSCD and document that CAOMECS grafting used to reconstruct corneal epithelium may reduce the expression levels of ALDH enzymes.

Corneal Collagen Crosslinking (CXL) for Corneal Ectasia after Small Incision Lenticule Extraction (SMILE). / Korneales Kollagen-Crosslinking (CXL) bei Hornhautektasie nach SMILE (Small Incision Lenticule Extraction).

With an estimated incidence of 0.011%, the SMILE procedure seems to have the lowest risk of postoperative keratectasia among contemporary keratorefractive procedures. Nevertheless, due to the novelty of the procedure as well as the lack of data, no clear superiority over femto-LASIK or PRK can be stated at this time. In this respect, application of the identical tomographic screening criteria previously developed for excimer-based procedures is of paramount importance to minimize the risk of corneal ectasia. As an adjunct to conventional corneal tomography, newer imaging modalities such as OCT-based epithelial mapping should be used for preoperative screening before keratorefractive surgery. Corneal crosslinking is an established treatment modality for post-SMILE keratectasia, which promises high success rates especially in early stages. The present case report illustrates these diagnostic and therapeutic considerations.

Integration and remodelling of a collagen anterior lamellar keratoplasty graft in an animal model - A preliminary report.

There is an international shortage of donor corneas for transplantation to treat the 1.5-2.0 million new cases of blindness secondary to corneal disease. Research has therefore been directed towards the development of artificial corneas using alternative materials such as collagen. The biocompatibility of an acellular collagen-based scaffold for anterior lamellar keratoplasty was investigated in vivo in a rabbit model. This scaffold has previously shown promise as a corneal substitute in vitro. Slit-lamp and Optical Coherence Tomography examinations were carried out at 2 weeks, 1, 2, 3, and 6 months post-operatively. Graft-host integration was investigated using immunohistochemistry of the cornea at 6 months. Results showed that the graft was biocompatible, supported corneal re-epithelialisation, and showed no signs of rejection. Migration of stromal cells into areas of the graft was observed, however this was accompanied by extensive graft digestion. Whilst the scaffold was biocompatible, further modifications to the material or supplementation with matrix metalloproteinase inhibitors are required to bring us closer to a stable and fully integrated corneal substitute.

Comparison of the rabbit and human corneal endothelial proteomes regarding proliferative capacity.

The shortage of human donor corneas has raised important concerns about engineering of corneal endothelial cells (CECs) for clinical use. However, due to the limited proliferative capacity of human CECs, driving them into proliferation and regeneration may be difficult. Unlike human CECs, rabbit CECs have a marked proliferative capacity. To clarify the potential reason for this difference, we analysed the proteomes of four human corneal endothelium samples and four rabbit corneal endothelium samples with quantitative label-free proteomics and downstream analysis. We discovered that vitamin and selenocompound metabolism and some signaling pathways such as NF-kappa B signaling pathway differed between the samples. Moreover, TGFß, PITX2 and keratocan were distinctively expressed in rabbit samples, which might be associated with active proliferation in rabbit CECs. This study illustrates the proteomic differences between human and rabbit CECs and might promote CEC engineering strategies.