Growth factors and mechano-regulated reciprocal crosstalk with extracellular matrix tune the keratocyte-fibroblast/myofibroblast transition.

Improper healing of the cornea after injury, infections or surgery can lead to corneal scar formation, which is associated with the transition of resident corneal keratocytes into activated fibroblasts and myofibroblasts (K-F/M). Myofibroblasts can create an extracellular matrix (ECM) niche in which fibrosis is promoted and perpetuated, resulting in progressive tissue opacification and vision loss. As a reversion back to quiescent keratocytes is essential to restore corneal transparency after injury, we characterized how growth factors with demonstrated profibrotic effects (PDGF, FGF, FBS, TGFß1) induce the K-F/M transition, and whether their withdrawal can revert it. Indeed, the upregulated expression of αSMA and the associated changes in cytoskeletal architecture correlated with increases in cell contractility, fibronectin (Fn) and collagen matrix density and Fn fiber strain, as revealed by 2D cell culture, nanopillar cellular force mapping and a FRET-labeled Fn tension probe. Substrate mechanosensing drove a more complete K-F/M transition reversal following growth factor withdrawal on nanopillar arrays than on planar glass substrates. Using decellularized ECM scaffolds, we demonstrated that the K-F/M transition was inhibited in keratocytes reseeded onto myofibroblast-assembled, and/or collagen-1-rich ECM. This supports the presence of a myofibroblast-derived ECM niche that contains cues favoring tissue homeostasis rather than fibrosis.

How Modifications of Corneal Cross-Linking Protocols Influence Corneal Resistance to Enzymatic Digestion and Treatment Depth.

Purpose: The purpose of this study was to determine the effects of the Photoactivated Chromophore for Keratitis Corneal Cross-Linking (PACK-CXL) protocol modifications on corneal resistance to enzymatic digestion and treatment depth. Methods: Eight hundred one ex vivo porcine eyes were randomly divided into groups of 12 to 86 corneas, treated with various epi-off PACK-CXL modifications, including acceleration (30 > 2 minutes, 5.4 J/cm2), increased fluence (5.4 > 32.4 J/cm2), deuterium oxide (D2O) supplementation, different carrier types (dextran versus hydroxypropyl methylcellulose [HPMC]), increased riboflavin concentration (0.1 > 0.4%), and riboflavin replenishment during irradiation (yes/no). Control group eyes did not receive PACK-CXL. A pepsin digestion assay was used to determine corneal resistance to enzymatic digestion. A phalloidin fluorescent imaging assay was used to determine the PACK-CXL treatment effect depth. Differences between groups were evaluated using a linear model and a derivative method, respectively. Results: PACK-CXL significantly increased corneal resistance to enzymatic digestion compared to no treatment (P < 0.03). When compared to a 10 minute, 5.4 J/cm2 PACK-CXL protocol, fluences of 16.2 J/cm2 and higher increased corneal resistance to enzymatic digestion by 1.5- to 2-fold (P < 0.001). Other protocol modifications did not significantly change corneal resistance. A 16.2 J/cm2 fluence also increased collagen compaction in the anterior stroma, whereas omitting riboflavin replenishment during irradiation increased PACK-CXL treatment depth. Conclusions: Increasing fluence will likely optimize PACK-CXL treatment effectiveness. Treatment acceleration reduces treatment duration without compromising effectiveness. Translational Relevance: The generated data help to optimize clinical PACK-CXL settings and direct future research efforts.

How the mechanobiology orchestrates the iterative and reciprocal ECM-cell cross-talk that drives microtissue growth.

Controlled tissue growth is essential for multicellular life and requires tight spatiotemporal control over cell proliferation and differentiation until reaching homeostasis. As cells synthesize and remodel extracellular matrix, tissue growth processes can only be understood if the reciprocal feedback between cells and their environment is revealed. Using de novo-grown microtissues, we identified crucial actors of the mechanoregulated events, which iteratively orchestrate a sharp transition from tissue growth to maturation, requiring a myofibroblast-to-fibroblast transition. Cellular decision-making occurs when fibronectin fiber tension switches from highly stretched to relaxed, and it requires the transiently up-regulated appearance of tenascin-C and tissue transglutaminase, matrix metalloprotease activity, as well as a switch from α5ß1 to α2ß1 integrin engagement and epidermal growth factor receptor signaling. As myofibroblasts are associated with wound healing and inflammatory or fibrotic diseases, crucial knowledge needed to advance regenerative strategies or to counter fibrosis and cancer progression has been gained.

Medical Management versus PACK-CXL in Dogs with Infectious Keratitis: A Randomized Controlled Trial Protocol.

Infectious keratitis is a common and painful disease, usually caused by bacteria in dogs. Brachycephalic breeds are at increased risk. Despite medical therapy, enzymatic corneal melting can lead to ulcer perforation and globe loss. Treatment alternatives are needed due to an increase in antibiotic resistance and growing popularity of brachycephalic dogs. Photoactivated Chromophore for Keratitis-Corneal Cross-linking (PACK-CXL) reduces enzymatic collagenolysis and damages multiple targets within microorganisms, resulting in corneal tissue stabilization and elimination of bacteria, irrespective of their antibiotic resistance status. A randomized controlled trial providing evidence of PACK-CXL effectiveness in dogs is lacking. We aim to determine whether PACK-CXL is a viable alternative to conventional medical therapy for canine infectious keratitis. Two hundred-and-seventy client-owned dogs with presumed infectious keratitis will be allocated to two equally sized treatment groups (PACK-CXL or medical therapy) in a masked, randomized, controlled, multicenter trial in eleven clinics. The primary outcome measure is treatment success defined as complete epithelial closure within 28 days. The sample size is based on a group sequential design with two interim analyses, which will be overseen by a Data Safety and Monitoring Board. Ethical approvals have been obtained. The study protocol is preregistered at preclinicaltrials.eu. Publishing trial protocols improves study reproducibility and reduces publication bias.

Outer retinal thickness and visibility of the choriocapillaris in four distinct retinal regions imaged with spectral domain optical coherence tomography in dogs and cats.

PURPOSE: To evaluate the outer retinal band thickness and choriocapillaris (CC) visibility in four distinct retinal regions in dogs and cats imaged with spectral domain optical coherence tomography (SD-OCT). To attempt delineation of a fovea-like region in canine and feline SD-OCT scans, aided by the identification of outer retinal thickness differences between retinal regions. METHODS: Spectralis® HRA + OCT SD-OCT scans from healthy, anesthetized dogs (n = 10) and cats (n = 12) were analyzed. Scanlines on which the CC was identifiable were counted and CC visibility was scored. Outer nuclear layer (ONL) thickness and the distances from external limiting membrane (ELM) to retinal pigment epithelium/Bruch's membrane complex (RPE/BM) and ELM to CC were measured in the area centralis (AC), a visually identified fovea-like region, and in regions superior and inferior to the optic nerve head (ONH). Measurements were analyzed using a multilevel regression. RESULTS: The CC was visible in over 90% of scanlines from dogs and cats. The ONL was consistently thinnest in the fovea-like region. The outer retina (ELM-RPE and ELM-CC) was thickest within the AC compared with superior and inferior to the ONH in dogs and cats (p < .001 for all comparisons). CONCLUSIONS: The CC appears a valid, albeit less than ideal outer retinal boundary marker in tapetal species. The AC can be objectively differentiated from the surrounding retina on SD-OCT images of dogs and cats; a fovea-like region was identified in dogs and its presence was suggested in cats. These findings allow targeted imaging and image evaluation of these regions of retinal specialization.

Penetrating Keratoplasty in Dogs using Acellular Porcine Corneal Stroma (BioCorneaVet™): A prospective pilot study of five cases.

OBJECTIVE: This prospective pilot study was conducted to evaluate the outcome of a commercially available corneal stroma substitute, Acellular Porcine Corneal Stroma (APCS), in dogs undergoing penetrating keratoplasty (PK) to restore corneal integrity after having deep ulcers. METHOD: Five dogs (1 eye in each dog) underwent a PK using APCS (BioCorneaVet™) as a graft. The surgical procedure and peri- and postoperative treatment were standardized. All cases required a minimum 6 months follow-up. Ease of keratoprosthetic tissue handling, graft survival, anterior chamber stability, corneal opacity, neovascularization and re-epithelialization were noted. Presence of secondary uveitis was investigated. RESULTS: BioCorneaVet™ was easy to handle and, at all-time points, provided adequate tectonic support. Graft survival was achieved in all 5 cases. A minimum follow-up period of 10 months was available for the five eyes (22 months maximum). Degree and area of corneal graft opacity progressively improved resulting in minimal to moderate loss of transparency in all cases but one, where it was severe. Neovascularization degree was most severe 0.5-1 month after surgery and fully resolved 4-6 months post-surgery. Re-epithelialization was complete in the majority of grafts in 1 month. Secondary uveitis was not detected at any time in 4 of 5 dogs. CONCLUSION: BioCorneaVet™ seems to be an effective graft for PK in the dog. In this case series, APCS was convenient to handle during surgery and provided excellent tectonic support. The material showed good tissue biocompatibility and resulted in the majority of cases in minimal to moderate graft opacity, that ameliorates with time.

Leptospiral antibody prevalence and surgical treatment outcome in horses with Equine Recurrent Uveitis (ERU) in Switzerland.

OBJECTIVE: To evaluate leptospiral antibody prevalence in 65 horses with ERU and compare outcome in 36 surgically treated eyes (2010-2015). PROCEDURES: Retrospective data analysis of horses with ERU (n = 65). C-value calculation with microagglutination assay titer (MAT) results for Leptospira spp. Evaluation of follow-up data after pars plana vitrectomy (PPV, n = 21 eyes) and suprachoroidal cyclosporine device implantation (SCDI, n = 15 eyes). Differences between groups were statistically analyzed using Fishers exact test, significance set at P < .05. RESULTS: Positive leptospiral titers were found in 28/65 blood, 31/65 aqueous humor (AH), and 19/20 vitreal (post-PPV) samples. The most common intraocular serovars were Leptospira interrogans grippotyphosa, pomona, and bratislava. Intraocular antibody production was suspected in samples of 22 horses (c-values > 1). Mean follow-up of surgical cases was 3.8 years (PPV) and 3.4 years (SCDI). PPV was performed in 21 eyes with positive, SCDI in 15 eyes with negative leptospiral test results. Uveitis recurred less often after PPV (2/21) compared to SCDI (6/15, P = .04). Retinal detachment occurred after PPV only (5/21, SCDI 0/15, P = .06), whereas only SCDI-treated eyes were enucleated (PPV 0/21, SCDI 3/15, P = .06). Blindness or visual impairment was equally likely to occur in both treatment groups after surgery (PPV 7/21, SCDI 7/15, P = .5). CONCLUSIONS: Leptospiral antibody prevalence is high in horses with ERU in Switzerland. Recurrence of uveitis is uncommon following PPV in the present study; an increased risk of retinal detachment exists. Enucleation is more often warranted in horses after SCDI in this study due to a higher uveitis recurrence.

Combined cSLO-OCT imaging as a tool in preclinical ocular toxicity testing: A comparison to standard in-vivo and pathology methods.

INTRODUCTION: Confocal scanning laser ophthalmoscopy and optical coherence tomography (cSLO-OCT) became available for human and animal ophthalmic examinations in recent years. The purpose of this study was to evaluate lesion detection and localization with cSLO-OCT imaging in an experimental outer retinal toxicity model and to compare cSLO-OCT to standard examination methods (indirect ophthalmoscopy (IO), fundus photography (FP) and central section histopathology). METHODS: A test compound was orally administered to albino rats (n = 4) for four weeks (part A) and to albino (n = 2) and pigmented (n = 2) rats for eight weeks (part B). Control animals received vehicle only. Retinal changes were documented using cSLO-OCT, IO, FP, angiography and histopathology. Retinal thicknesses were compared between groups using a mixed effects model. RESULTS: All compound-treated animals developed progressive multifocal hyperreflective spot changes mostly confined to the retinal pigment epithelium. In study parts A and B, cSLO identified fundus lesions earlier than IO/FP in albino rats. In study part B, cSLO quantified fundus lesions more accurately than IO/FP in albino rats but no difference was seen in pigmented rats. Central section histopathology revealed no abnormalities in three out of four compound-treated animals in part B. Altogether, without cSLO-OCT, present fundus changes would have remained undetected in one of four compound-treated animals in both parts A and B. DISCUSSION: Integration of combined cSLO-OCT imaging into toxicology study design can improve toxicity study readouts and facilitate longitudinal examination of single animals at multiple time points, leading to a reduction of experimental animal numbers.

Corneal cross-linking as a treatment for corneal dystrophy with secondary bacterial infection in a Friesian horse.

Corneal cross-linking should be considered as treatment option in Friesian horses with infectious keratitis and corneal dystrophy. Optical coherence tomography, giving information of corneal structure, can help for diagnosis and monitoring.

Ocular and periocular radiation toxicity in dogs treated for sinonasal tumors: A critical review.

Visual impairment from radiation-induced damage can be painful, disabling, and reduces the patient's quality of life. Ocular tissue damage can result from the proximity of ocular organs at risk to irradiated sinonasal target volumes. As toxicity depends on the radiation dose delivered to a certain volume, dose-volume constraints for organs at risk should ideally be known during treatment planning in order to reduce toxicity. Herein, we summarize published ocular toxicity data of dogs irradiated for sinonasal tumors from 36 publications (1976-2018). In particular, we tried to extract a dose guideline for a clinically acceptable rate of ocular toxicity. The side effects to ocular and periocular tissues were reported in 26/36 studies (72%) and graded according to scoring systems (10/26; 39%). With most scoring systems, however, toxicities of different ocular and periocular tissues are summed into one score. Further, the scores were mostly applied in retrospect and lack volume- and dose-data. This incomplete information reflects the crux of the matter for radiation dose tolerance in canine ocular tissues: The published information of the last three decades does not allow formulating dose-volume guidelines. As a start, we can only state that a mean dose of 39 Gy (given in 10 x 4.2 Gy fractions) will lead to loss of vision by one or both eyes, while mean doses of <30 Gy seem to preserve functionality. With a future goal to define tolerated doses and volumes of ocular and periocular tissues at risk, we propose the use of combined ocular toxicity scoring systems.

Evaluation of D-dimer levels in aqueous humor of rabbit eyes with and without induced intraocular fibrin and fibrinolytic treatment.

OBJECTIVE: To analyze D-dimer concentrations in aqueous humor (AH) of rabbit eyes under physiological conditions, after induction of fibrin clots, and following fibrinolytic therapy. ANIMALS STUDIED: Prospective study measuring D-dimers in aqueous humor of rabbit eyes with induced fibrin clots (n = 44). PROCEDURES: Rabbits were purchased in two groups, which led to two temporally separated experimentation groups. Different treatment protocols were compared for their efficacy in fibrin reduction (slit-lamp examination, high-resolution ultrasound). AH was taken from left eyes before clot induction (baseline, day 1), 24 hours later after clot establishment/prior to drug administration (post-induction, day 2) and 48 hours after clot induction (post-treatment, day 3). An enzyme-linked immunosorbent assay (ELISA) was performed to measure intraocular D-dimer concentrations RESULTS: D-dimer concentrations were measurable in all samples. There were no differences in D-dimer levels across time points or treatments within the arrival groups. However, a significant difference in mean D-dimer levels was observed between the two arrival groups (group 1:3.1 µg/mL; group 2:6.1 µg/mL; P < .0001), which made a direct comparison of treatment groups impossible. Clinically, all eyes displayed fibrin clots in the anterior chamber and different treatment types led to significant differences in clot resolution (clot size reduction after intracameral treatment: 98%, topical treatment: 60%, no treatment: 40%). CONCLUSION: D-dimers were identified in all AH samples of rabbits with large variability between samples. D-dimer levels were neither predictive for differences in induced fibrin formation nor for drug efficacy.

The localization of a conjunctivoscleral foreign body via high-resolution microscopy coil magnetic resonance imaging in a dog.

A 3-year-old French bulldog was presented to the ophthalmology service of the Vetsuisse Faculty, University of Zurich with a 3-day history of conjunctival swelling of the left eye (OS). Ophthalmologic examination revealed a moderate conjunctival hyperemia and chemosis. A migrating foreign body having entered the conjunctival fornix behind the nictitating membrane was suspected. Within the first 24 hours of medical management, OS developed a panuveitis and a scleral perforation was highly suspected. Ocular and orbital ultrasound as well as conventional magnetic resonance imaging (MRI) examinations failed to confirm the presence of a perforating foreign body. A High-Resolution MRI (HR-MRI) using a microscopy coil was then performed with findings consistent with a perforating and migrating foreign body. A grass awn of 12 mm length was surgically retrieved "ab externo" from its' point of entry into the sclera. To the best of our knowledge, HR-MRI has not yet been used to examine canine eyes. This case report supports the idea that orbital imaging can be greatly enhanced with the introduction of HR-MRI using microscopy coils with clinically relevant implications.

Seasonal Effects on the Corneoconjunctival Microflora in a Population of Persian Cats in Iran.

This study was performed to determine the normal seasonal aerobic and an-aerobic corneoconjunctival bacterial flora in cats. Thirty eyes of 15 clinically normal client-owned Persian cats were evaluated. All cats lived in a similar indoor/outdoor home environment being fed the same diet for the entire year. The cats did not receive any medications and were found to be clinically healthy 1 week prior to each microbial sampling. The cats were not exposed to other cats during the study period. Microbial samples were collected at the same time of day on the first day of the second month of each of the four seasons. During sample collection, a sterile swab was rolled over the corneoconjunctival surface avoiding contact with surrounding skin or hair. Immediately after sample collection, microbiologic aerobic and anaerobic cultures were initiated. Gram-positive bacteria were the most prevalent isolates. The most commonly isolated bacterial organisms across all seasons were Staphylococcus epidermidis (41/95; 43.2%), ß-hemolytic streptococcus (18/95; 18.9%), Staphylococcus aureus (17/95; 17.9%), and Escherichia coli (11/95; 11.5%). Twenty-five cultures of a total of 120 (20.8%) were negative. One negative culture was collected in the summer, while 21 cultures were negative in fall and winter. Gram-positive bacteria were the predominant micro-organisms of the normal ocular surface of healthy cats in all seasons in this study. This result is in agreement with previous publications.

Protective coatings for intraocular wirelessly controlled microrobots for implantation: Corrosion, cell culture, and in vivo animal tests.

Diseases in the ocular posterior segment are a leading cause of blindness. The surgical skills required to treat them are at the limits of human manipulation ability, and involve the risk of permanent retinal damage. Instrument tethering and design limit accessibility within the eye. Wireless microrobots suturelessly injected into the posterior segment, steered using magnetic manipulation are proposed for procedures involving implantation. Biocompatibility is a prerequisite for these procedures. This article investigates the use of polypyrrole- and gold-coated cobalt-nickel microrobots. While gold has been used in ocular implants, no ocular implantation involving polypyrrole is reported, despite its well-established biocompatibility properties. Coated and uncoated microrobots were investigated for their corrosion properties, and solutions that had contained coated and uncoated microrobots for one week were tested for cytotoxicity by monitoring NIH3T3 cell viability. None of the microrobots showed significant corrosion currents and corrosion potentials were as expected in relation to the intrinsic nobility of the materials. NIH3T3 cell viability was not affected by the release medium, in which coated/uncoated microrobots were stored. In vivo tests inside rabbit eyes were performed using coated microrobots. There were no significant inflammatory responses during the first week after injection. An inflammatory response detected after 2 weeks was likely due to a lack of longer-duration biocompatibility. The results provide valuable information for those who work on implant technology and biocompatibility. Coated microrobots have the potential to facilitate a new generation of surgical treatments, diagnostics and drug-delivery techniques, when implantation in the ocular posterior segment will be possible. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 836-845, 2017.

Sub-Tenon's injection in equine cadaver eyes: MRI visualization of anesthetic fluid distribution and comparison of two different volumes.

OBJECTIVE: To compare the localization and distribution of two different anesthetic fluid volumes around equine cadaver eyes to determine an appropriate volume for a single sub-Tenon's injection in horses. PROCEDURE: A single sub-Tenon's injection of 2% lidocaine was performed in 10 equine cadaver heads (20 eyes) using two different volumes (7 mL on one side and 10 mL on the opposite side). The posterior circular distribution of the anesthetic was quantified in sagittal, dorsal, and transverse MRI (T2W-TSE) sequences and evaluated independently by three board-certified radiologists. The distribution of the two fluid volumes was compared via a paired Student's t-test. The interobserver reliability was evaluated via a Kruskal-Wallis test. RESULTS: Extension of the injection fluid was observed along the dorsal and temporal quadrants of the globe within the subconjunctival space, the anterior and posterior sub-Tenon's space, and into the muscle sheaths along the extraocular muscles. Accumulation of anesthetic fluid directly surrounding the optic nerve was detected in three of 20 cadaver eyes. Circular distribution of the 7 and 10 mL anesthetic volumes was not significantly different (P = 0.849). More retrograde leakage of the anesthetic was observed using the 10 mL volume. Evaluation of interobserver reliability revealed no significant differences between observers (P = 0.21-0.92). CONCLUSIONS: Sub-Tenon's anesthesia can have potential as an alternative to retrobulbar anesthesia for ophthalmic surgeries in equines. A 7- to 10-mL injection volume should be appropriate based on the results of this study. The distribution of the anesthetic solution in live tissues, the clinical effects, and the potential for complications will have to be evaluated in vivo.

Penetration depth of corneal cross-linking with riboflavin and UV-A (CXL) in horses and rabbits.

OBJECTIVE: CXL penetration depth is an important variable influencing clinical treatment effect and safety. The purposes of this study were to determine the penetration depth of CXL in rabbit and equine corneas in epithelium-on and epithelium-off procedures and to assess an ex vivo fluorescent biomarker staining assay for objective assessment of CXL penetration depth. PROCEDURES: CXL treatment was performed according to a standardized protocol on 21 and 17 rabbit eyes and on 12 and 10 equine eyes with and without debridement, respectively. Control corneas were treated similarly, but not exposed to CXL. Hemicorneas were stained with either phalloidin and DAPI to visualize intracellular F-actin and nuclei, or with hematoxylin and eosin. Loss of actin staining was measured and compared between groups. RESULTS: Epithelium-off CXL caused a median actin cytoskeleton loss with a demarcation at 274 µm in rabbits and 173 µm in horses. In non-CXL-treated controls, we observed a median actin cytoskeleton loss with a demarcation at 134 µm in rabbits and 149 µm in horses. No effect was detected in the epithelium-on procedure. CONCLUSIONS: CXL penetration depth, as determined by a novel ex vivo fluorescent assay, shows clear differences between species. A distinct effect was observed following epithelium-off CXL treatment in the anterior stroma of rabbits, but no different effect was observed in horses in comparison with nontreated controls. Different protocols need to be established to effectively treat equine patients with infectious corneal disease.

Treatment of bullous keratopathy with corneal collagen cross-linking in two dogs.

OBJECTIVE: Corneal collagen cross-linking with riboflavin and UV-A (CXL) decreases corneal oedema and increases visual acuity in human patients with bullous keratopathy. Presumed mechanisms are an increase in collagen packing density and a reduction in stromal swelling pressure. We present two cases in which CXL was used to treat bullous keratopathy in dogs. PROCEDURES: Four eyes of two dogs with painful bullous keratopathy-induced corneal erosions that were resistant to prior therapy were treated with CXL. Both corneas of the second patient were dehydrated to ± 400 µm corneal thickness using topical 70% glycerol solution immediately prior to CXL. Follow-up included slit-lamp examination, fluorescein staining and photographic documentation in both cases and high-resolution ultrasound examination in the second patient. RESULTS: All four eyes were comfortable and fluorescein negative at 1-week post-CXL and remained so for the rest of the follow-up period (17.5 months for case 1 and 6 months for case 2). The owner of the first patient reported a less oedematous cornea and improvement in vision that lasted for 6 months. Despite a reported lack of improvement in vision in the second patient, corneal thickness initially decreased, but was back at baseline thickness at the 4-month recheck. CONCLUSIONS: Similar to humans, CXL might become a useful treatment option for bullous keratopathy-induced therapy-resistant corneal erosions in dogs. Patient comfort was greatly improved, but corneal thickness decrease was not as long-lasting as reported for humans. The presently used protocols might need modification to fit the dog cornea.

Corneal collagen cross-linking (CXL) for the treatment of melting keratitis in cats and dogs: a pilot study.

OBJECTIVE: UV-A/riboflavin cross-linking (CXL) of corneal collagen fibers is an established, highly promising therapy for corneal melting in physician-based ophthalmology. A prospective pilot study was conducted to demonstrate proof of principle of this novel method for the treatment of melting corneal ulcers in dogs and cats. PROCEDURES: After obtaining owner consent, CXL was performed in three cats and three dogs with corneal melting, which either affected the entire corneal surface or was resistant to conventional antibiotic and anticollagenolytic therapy, and affected parts or all of the corneal surface. Medical therapy was continued in all patients. The available follow-up ranged from 2 to 22.5 months and involved slit-lamp examination, fluorescein staining, and photographic documentation during all rechecks. RESULTS: Surgical stabilization of the cornea was not necessary in any case, because progression of corneal melting was arrested in all cases within 1-20 days of CXL treatment. Corneal re-epithelization occurred within 7-40 days in all eyes. At 40 days after CXL, all eyes presented a quiescent corneal state without signs of active inflammation and with beginning scar formation. The complications observed in three of the six animals included a corneal sequestrum, superficial corneal stromal pigmentation, and bullous keratopathy. CONCLUSIONS: This study shows the feasibility of CXL to treat progressive corneal melting in veterinary patients. CXL may represent a cost-efficient and safe alternative therapy in the treatment for corneal melting in veterinary ophthalmology. More investigations comparing the effectivity and complication rate of CXL to those of standard medical treatment are necessary.

Corneal collagen cross-linking as treatment for infectious and noninfectious corneal melting in cats and dogs: results of a prospective, nonrandomized, controlled trial.

OBJECTIVE: UV-A/Riboflavin cross-linking of corneal collagen fibers (CXL) is a highly promising therapy for corneal melting in humans. A prospective interventional, nonrandomized, controlled study was conducted to compare the stabilizing effect of CXL treatment on melting keratitis in dogs and cats and the complication rate of CXL to those of standardized intensive medical treatment. PROCEDURES: Forty-nine eyes with melting keratitis were included in the study between October 2009 and October 2012. All eyes were treated according to the same medical treatment protocol. Nineteen eyes were CXL-treated, and 30 eyes were not. Follow-up included slit-lamp examination, fluorescein staining, ulcer size measurement, stromal stability evaluation, photographic documentation, and documentation of complications. RESULTS: Five of 19 eyes in the CXL group and 9/30 eyes in the control group required rescue stabilization due to continued melting. Seven of the nine control group corneas stabilized after rescue CXL treatment. At initial presentation, the ulcers in the canine CXL group were significantly deeper and larger than in the control group. Ulcer deepening during follow-up was more pronounced in the canine control group than in the canine CXL group. CXL treatment-related complications were not observed. CONCLUSIONS: Based on the similar failure rates in the control and CXL treatment groups despite the poorer initial situation in the CXL group, the tendency for the ulcers in the control group to deepen and the stabilization of all corneas receiving CXL rescue treatment, we believe that CXL has its place as an adjunctive therapy for melting keratitis in veterinary ophthalmology.

Evaluation of the conjunctival fungal flora and its susceptibility to antifungal agents in healthy horses in Switzerland.

OBJECTIVE: To characterize the conjunctival fungal flora and to determine the susceptibility of 2 isolated molds to antifungal drugs in samples of 64 healthy horses from The National Stud in Switzerland. PROCEDURE: Conjunctival cytobrush samples were collected from both eyes of 64 ophthalmologically normal horses in August 2012 and subsequently cultured on Sabouraud's agar medium. Growing fungi were identified and counted. Etests or broth microdilution tests for Aspergillus fumigatus and Eurotium amstelodami were carried out to determine antifungal drug sensitivity. These species had previously been detected in samples from eyes with keratomycosis in Switzerland. Minimal inhibitory concentrations (MICs) for voriconazole, fluconazole, itraconazole, amphotericin B, and miconazole were recorded. RESULTS: Fifty-nine of the horses were tested positive for fungal growth from at least one eye (92%). Eleven genera of fungi were identified. The most common fungal genera were Alternaria, Eurotium, Rhizopus, and Cladosporium. Aspergillus spp. and Penicillium spp. were isolated frequently, while no Fusarium spp. was found. In only 2 cases, yeasts were identified as Candida guilliermondii. For certain fungal species, the type of bedding and housing appeared to influence their prevalence. Susceptibility testing of A. fumigatus showed lowest MICs for voriconazole, E. amstelodami for voriconazole and itraconazole. High MICs for fluconazole were detected for all tested fungi while MICs for amphotericin B and miconazole were variable. CONCLUSIONS: A large range of fungal mold species was identified including A. fumigatus and E. amstelodami, which have been causative agents of keratomycosis in Switzerland. Best in vitro susceptibility results for these two species were obtained for voriconazole.