PURPOSE:To investigate the presence and distribution of l-kynurenine aminotransferases immunoreactivity in human and animal lenses during cataract formation.METHODS:Immunohistochemistry was conducted using polyclonal antibodies against KAT I, KAT II and KAT III on sections of 26 anterior capsules from patients undergoing surgical treatment of anterior subcapsular cataract (ASC) and 22 cataractous lenses from human eyes enucleated because of choroidal malignant melanoma. Additionally, the eyes of 11-month-old DBA/2J mice (6 eyes) were investigated (with KAT I and II). Ten clear human lenses and four BL6 mice lenses were used as controls. Spatial immunoreactivity patterns of enzymes were compared with Periodic Acid - Schiff (PAS)-stained sections.RESULTS:Immunohistochemical analysis revealed presence of KAT I, KAT II and KAT III in extracellular structures of all studied types of cataract in human eyes showing specific pattern of the stain. In cortical cataract, immunoreactivity was observed on cortical lens fibres. In nuclear cataract, KAT II revealed stronger and diffused staining than KAT I. Additionally, both KAT showed more pronounced staining at the edge of small clefts. In normal human lenses, KAT I, II and III, immunoreactivity was not observed. Presence of KAT I and KAT II in the intercellular substance of DBA/2J mice cataract was observed. In BL6 mice lenses without cataract, only weak KAT I and KAT II staining was observed.CONCLUSIONS:Presence of l-kynurenine aminotransferases in extracellular matrix (ECM) during human cataract formation suggests that products of l-kynurenine pathway might be involved in mechanisms of cataractogenesis.

PURPOSE:During deep anterior lamellar keratoplasty (DALK), endothelium and Descemet's membrane are separated from the corneal stroma by intrastromal air injection ('big-bubble technique'). The aim of our study is to analyse histopathological changes in host corneal tissue caused by air insufflation in patients with keratoconus, their variability in 10 patients and their possible clinical implication.METHODS:The excised anterior corneal lamellae of 10 patients with keratoconus having undergone DALK using the 'big-bubble technique' were analysed by light and transmission electron microscopy as well as immunohistochemistry. In addition, intrastromal air accumulations were quantified morphometrically.RESULTS:Intrastromal air was detected in all examined excised lamellae (8% of stromal volume), but with large variability (SD 8.8). It was detected preferentially in the inner layer of the corneal stroma and represented there up to 39% of the stromal volume. In addition, the air was predominantly located at one periphery of the excised lamellae. Intrastromal air bubbles were larger in the inner than in the superficial stromal layer and characterized by round shape and a CD68-negative collagenous 'pseudocapsule'. We detected no air-injection-induced alterations in Bowman's layer and epithelium.CONCLUSION:Our results show that 'big-bubble DALK' causes significant intrastromal air accumulations in the cornea. Pathologists should be conscious of this phenomenon and the high topographic variability. Intrastromal air in the recipient rim may be accompanied by a decrease in mechanical stability and could contribute to postoperative suture loosening.

Background: This retrospective study examines the histopathological changes, especially the occurrence of retrocorneal membranes, in irreversible graft failure after penetrating keratoplasty.Patients/Materials and Methods: 371 corneas of 308 patients were examined. The examination was carried out using a light microscope.Results: 45% of the corneas (167/371) showed a retrocorneal membrane with a thickness of 2-520 micrometers. Re-endothelialisation was detected in 75 cases. In 74% (124/167) cellular infiltration into the stroma could be observed. In 32% (119/371) the graft-host border was visible.Conclusions: Retrocorneal membranes are a frequent finding in irreversible graft failure after penetrating keratoplasty. Aetiologically the graft-host border as well as the formation of connective tissue seem to play a key role.

BACKGROUND:This retrospective study examines the histopathological changes, especially the occurrence of retrocorneal membranes, in irreversible graft failure after penetrating keratoplasty.PATIENTS/MATERIALS AND METHODS:371 corneas of 308 patients were examined. The examination was carried out using a light microscope.RESULTS:45% of the corneas (167/371) showed a retrocorneal membrane with a thickness of 2-520 micrometers. Re-endothelialisation was detected in 75 cases. In 74% (124/167) cellular infiltration into the stroma could be observed. In 32% (119/371) the graft-host border was visible.CONCLUSIONS:Retrocorneal membranes are a frequent finding in irreversible graft failure after penetrating keratoplasty. Aetiologically the graft-host border as well as the formation of connective tissue seem to play a key role.

PURPOSE:Bevacizumab eyedrops inhibit corneal neovascularization. The purpose of this study was to analyze the safety profile of VEGF-A neutralization at the ocular surface.METHODS:Bevacizumab eyedrops (5 mg/mL) and an antimurine VEGF-A antibody (250 microg/mL) were applied to normal murine corneas five times a day for 7 and 14 days. Subsequently, corneas were analyzed for morphologic changes by light and electron microscopy. In a mouse model of corneal epithelial abrasion, the effects of topically applied anti-VEGF antibodies on epithelial wound healing were analyzed: the treatment group received bevacizumab (5 mg/mL) or the antimurine VEGF-A antibody (250 microg/mL) as eyedrops, and the control group received an equal volume of saline solution. After 12, 18, and 24 hours, corneas were photographed in vivo with and without fluorescein staining for morphometry. Afterwards the mice were killed, and eyes were removed for histology, immunohistochemistry with Ki67/DAPI, and electron microscopy. The effect of midterm anti-VEGF therapy on corneal nerve density was assessed by staining corneas treated with an FITC-conjugated anti-neurofilament antibody and morphometric analysis.RESULTS:Murine corneas treated with two different types of anti-VEGF antibody eyedrops did not show obvious corneal morphologic changes at the light and electron microscopic levels. Furthermore, anti-VEGF antibody eyedrops had no significant impact on the wound healing process after corneal epithelial injury or on normal murine corneal nerve fiber density.CONCLUSIONS:Topical neutralization of VEGF-A at the corneal surface does not have significant side effects on normal corneal epithelial wound healing, normal corneal integrity, or normal nerve fiber density. Therefore, anti-VEGF eyedrops seem to be a relatively safe option to treat corneal neovascularization.

Aim: To investigate the effect of timolol and latanoprost on the extracellular matrix organisation, inflammatory infiltration, and expression of matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinases (TIMPs) in the human conjunctiva.Methods: Conjunctival biopsies were obtained from the inferior fornix during routine cataract surgery from 20 patients with primary open-angle glaucoma, who had received a monotherapy either with timolol or latanoprost, and from 10 non-glaucomatous patients. Specimens were investigated by light microscopy, immunohistochemistry using antibodies against MMP-1,-3, TIMP-2,-3 and CD 68 antibodies and by quantitative transmission electron microscopy.Results: The number of collagen fibres was significantly decreased in latanoprost-treated conjunctival specimens compared with timolol-treated eyes (p<0.01) but showed no difference to controls. Amorphous material was increased in both treated groups compared with controls (p<0.001) but was less in latanoprost-treated specimens compared with timolol-treated eyes (p<0.001). Optically clear spaces, probably containing glycosaminoglycans, were significantly reduced in both treated groups-with less of a reduction in latanoprost compared with timolol-treated eyes (p<0.001). A marked upregulation of MMP-1 and MMP-3 and moderately increased staining for TIMP-2 and TIMP-3 was found in epithelial cells and subepithelial stromal cells of latanoprost-treated eyes. A moderate infiltration with macrophages and inflammatory cells was observed in timolol-treated eyes.Conclusions: Latanoprost-treated conjunctival specimens showed a decreased stromal collagen density and a less pronounced inflammatory infiltration. The upregulation of MMP-1 and MMP-3 in latanoprost-treated eyes might explain the reduced extracellular matrix accumulation in the conjunctival stroma. Therefore, latanoprost therapy might have a more favourable effect on the outcome of glaucoma filtering surgery.

Purpose DBA/2J mouse develop spontaneously changes in anterior chamber like pigment dispersion syndrome, iris atrophy, posterior synechiae leading to IOP increase. Additionally, there are observations suggesting cataract formation. The aim of this study was to describe features of cataract formation in DBA/2J mice. Moreover, presence and pattern of the L- kynurenine aminotransferases (KAT I , II and III) immunoreactivity in the cataractous lens of DBA/2J mice was investigated. Methods Immunohistochemistry was conducted using polyclonal antibodies against KAT I, II and KAT III on 18 cataractous lenses of 8-, 11- and 24-month old DBA/2J mice (6 in each group). All the sections of the lens were stained with Periodic Acid – Schiff (PAS), and normal anatomy of the anterior segment of the eye was confirmed. Moreover, all the animals subjected to the study were examined using slit lamp and anterior chamber photography. Results As observed clinically, the animals developed cataract during aging. PAS staining revealed presence of lens opacification. Immunohistochemical analysis revealed presence of KAT I, II and KAT III in the extracellular structures of cataract showing specific pattern of the stain. In cortical cataract the immunoreactivity was observed on cortical lens fibres. In nuclear cataract, KAT II revealed stronger and diffused staining than KAT I. Additionally both KATs showed more pronounced staining at the edge of small clefts. Conclusion Manifestation of L-kynurenine aminotransferases in extracellular matrix during cataract formation in DBA/2J mice suggests that products of L-kynurenine pathway might be involved in mechanisms of cataractogenesis in this animal model.

PURPOSE: To evaluate the efficacy of the laser photolysis system (LPS) (A.R.C. Laser GmbH) in removing lens epithelial cells (LECs) to prevent posterior capsule opacification (PCO) in an in situ model.SETTING: Department of Ophthalmology, Friedrich-Alexander-UIniversity Erlangen-Nurnberg, Erlangen, Germany.METHODS: Twelve enucleated porcine eyes fixed in a specially developed eye holder were randomly assigned to the control or treatment group. The cornea and iris were removed from all eyes, and a small paracentral capsulorhexis was performed. The lens nucleus and cortex were extracted by hydroexpression. The tip of the LPS was inserted into the capsular bag of eyes in the treatment group, and 50 pulses (10 mJ) were applied to the anterior capsule. All capsules were evaluated for remaining LECs by confocal laser scanning microscopy (HRT II with the Rostock Cornea Module, Heidelberg Engineering) and standard histology (hematoxylin-eosin and periodic acid-Schiff stains).RESULTS: In the control group, a homogenous layer of LECs attached to the anterior capsule was seen with both evaluation methods. In the treatment group, no LECs adherent to the anterior capsule were detected, suggesting complete ablation of LECs from the capsule. Small islands of equatorial LECs were found in places in which the remaining cortical fibers protected cells from the laser shockwave. The results of the confocal laser scanning microscopy were confirmed by standard histology.CONCLUSIONS: The LPS completely ablated LECs in an in situ model of cataract extraction. This system might prevent formation of PCO in vivo.

Ziel: In einer kürzlich am enukleierten Schweineauge durchgeführten In-situ-Studie konnten wir zeigen, dass eine vollständige Ablation von Linsenepithelzellen von der Linsenkapsel nach erfolgreicher Kataraktextraktion mithilfe des ARC-Laser-Photolyse-Systems (ARC-LPS) möglich ist, ohne die Linsenkapsel dabei zu gefährden. Ziel dieser Studie war die Untersuchung des Systems als Methode einer intraoperativen Nachstarprophylaxe in einem In-vivo-Modell am Kaninchenauge. Methode: Eine Gruppe von 12 New Zealand White Kaninchen wurde an beiden Augen einer Standardkatarakt-Operation unterzogen und in eine Kontroll- und Untersuchungsgruppe eingeteilt. Vor Implantation einer Faltlinse in den Kapselsack, wurde in die Augen der Untersuchungsgruppe das Handstück des ARC-LPS eingeführt und eine mittlere Pulszahl von 204 (±36) Pulsen mit einer Energie von je 7 mJ auf die verbliebene vordere und äquatoriale Linsenkapsel in mäanderförmigem Verteilungsmuster appliziert. Zur Evaluierung wurden 10 Augen nach 6 Wochen enukleiert, zwischen Limbus und Äquator eröffnet und digitale posteriore Bilder nach einer modifizierten Miyake/Apple-Technik angefertigt, um anschließend mithilfe der EPCO-Software planimetrisch im Bereich der Optik vermessen zu werden. Im Anschluss wurden Standardhistologiepräparate zur Evaluierung angefertigt. Ergebnisse: Die Evaluation der digitalen Miyake-Bilder mithilfe der EPCO-Software ergab bei den Augen der Untersuchungsgruppe im Vergleich zur Kontrollgruppe eine deutliche Reduktion des EPCO-Scores bzw. der Nachstarbildung: Kontrollgruppe=0,69, Untersuchungsgruppe=0,33. (p= 0,035). Intraoperativ kam es bei 2 von 8 Augen zu geringen Blutungen aus Irisgefäßen. Schlussfolgerungen: Die Laserablation mithilfe des ARC-LPS als intraoperative Nachstarprophylaxe führt nach sechswöchigem Beobachtungszeitraum zu einer wirksamen Reduktion des Nachstars. Mögliche Iritationen der Iris können durch Reduktion der aufgewandten Laserenergie minimiert werden, so dass dieses System zukünftig im Rahmen der Kataraktchirurgie beim Menschen eine intraoperative Linsenkapselpolitur mit Verminderung des Nachstarrisikos ermöglichen sollte.

Background: Corpora amylacea (CAm) are a hallmark of aging and neurodegeneration. The presence of kynurenine aminotransferases I and II (KAT I and II) in CAm in the human retina and optic nerve has been already shown. The present study aimed to examine kynurenine aminotransferase III (KAT III) immunoreactivity in CAm in the human retina and optic nerve.Material and methods: Polyclonal antibody against KAT III was used on sections of human eyes enucleated due to malignant uveal melanoma. PAS-stained sections of CAm were compared with KAT III stained ones.Results: KAT III immunoreactivity was observed in CAm in the retina, prelaminar, laminar and retrolaminar region of the optic nerve with similar location to PAS-stained sections. The most intense staining was observed in the retrolaminar part of the optic nerve. KAT III immunoreactivity was also present in the cytoplasm of retinal ganglion cells.Conclusions: Expression of KAT III in CAm in the human retina and optic nerve indicates that this enzyme may be relevant in mechanisms of neurodegeneration leading to CAm formation.

We saw a 62-year-old male with a conjunctival chemosis, which was resistant to local therapy. Excisional biopsy and histological work-up of the tissue were performed. Additional immunohistochemical staining using the Lyve-1 antibody (specific for lymphatic vascular endothelium) allowed the diagnosis of a lymph-angioma of the conjunctiva to be made