Intraocular Neurografts as a Model for Studying of Organization of Synaptic Connections in a Denervated Brain Area.

Intraocular neurografts of the septal region of rats were used as the model of deafferentiated brain area where the lack of adequate innervation is compensated for own interneuronal connections. Septum anlage from the brain of a 17-day fetus served as the donor material. The grafts developing in the anterior eye chamber over 3 months represented well-differentiated samples of the nervous tissue. A comparative morphometric study of the tripartite organization of synapses in the grafts and in the septum in situ was conducted. In the grafts, the mean volume and perimeter of synaptic terminals were below the normal. At the same time, postsynaptic densities did not differ from the control. A significant difference was found in the degree of surrounding of presynaptic terminals by astrocytic processes: in the grafts this parameter was higher by 1.8 times. Our results attest to an important role of perisynaptic glia in the formation of functionally active synaptic contacts with unusual neuronal targets.

A magnetic bead-based method for mouse trabecular meshwork cell isolation.

PURPOSE: Mice have been used widely for glaucoma research. However, due to the small size of the mouse eye, it is difficult to dissect mouse trabecular meshwork (MTM) tissues and establish MTM cell strains. To circumvent this problem, we took advantage of the phagocytic property of trabecular meshwork (TM) cells, and developed a novel magnetic bead-based method that enables us to isolate pure MTM cells. METHODS: After anesthesia, up to 2 µL of fluorescent or magnetic microbeads were injected intracamerally into the mouse eyes. To study the distribution and localization of the beads, mice were sacrificed 1 to 7 days after injection, and eyes were enucleated for fluorescent or transmission electron microscopy (TEM) study, respectively. To isolate MTM cells, anterior segments injected with magnetic beads were dissected from 10 to 15 sterilized mouse eyes 7 days after injection. The tissues were digested with collagenase A and purified by using a magnetic field as well as repeated washing. RESULTS: TEM studies showed that the magnetic beads were located in the mouse TM, but not in corneal or scleral fibroblast cells. Cultured MTM cells were similar morphologically to human TM cells. MTM cells expressed TM markers, including collagen IV, laminin, and α-smooth muscle actin. Also, MTM cells treated with 100 nM dexamethasone showed increased formation of cross-linked actin networks and induction of myocilin expression. CONCLUSIONS: The magnetic bead-based method is efficient for isolating MTM cells with minimal microdissection techniques required. It will be a useful approach for isolating TM cells from small animals for glaucoma research.

Intravitreal triamcinolone acetonide induced changes in the anterior segment in a pig model of branch retinal vein occlusion.

BACKGROUND: Triamcinolone acetonide (TA) has applications for the treatment of a large range of intraocular vascular diseases. The present study in pigs was performed to investigate histopathological and histochemical changes in the levels of myocilin deposition in the anterior segment in a model of branch retinal vein occlusion (BRVO) after vitreal administration of TA. METHODS: After ophthalmoscopic examination, intraocular pressure (IOP) measurement and fundus photography, a BRVO was created photothrombotically in each eye of six pigs, using argon green photocoagulation. The left eye was then injected intravitreally with 4 mg/0.1 ml TA. After 11 weeks, the eyes were re-examined, animals sacrificed, and eyes enucleated and processed in paraffin and epoxy resin. Immunofluorescence cytochemistry on paraffin sections was performed to localise the distribution of myocilin in the anterior segment and histology by light and transmission electron microscopy on epoxy resin sections on TA-treated and untreated eyes. RESULTS: Histology revealed pathological changes in the TA-treated eye, including swollen mitochondria, layered long endoplasmic reticulum, pleomorphic nuclei, dense fibrillar extracelluar deposits and aggregates of unusual cell inclusions. Myocilin levels were significantly higher in the TA-treated eyes in the trabecular meshwork (p = 0.001), ciliary process (p = 0.011) and iris (p = 0.030) than in the untreated eyes. CONCLUSIONS: This study suggests that increased myocilin synthesis and related ultrastructural changes in the anterior segment after treatment with intravitreal TA in a porcine model of retinal oedema in BRVO may contribute to IOP elevation.

[Possibilities of using long-wave irradiation in the near-infrared and red spectral ranges for visualization of eyeball structures].

The proposed study technique makes it possible to non-invasively visualize the structures of the anterior and posterior portions of the eyeball in the red and near-infrared spectral ranges of irradiation. Further study of the capacities of the technique and its introduction into clinical practice should be made.

Effect of the age cross-link breaker alagebrium on anterior segment physiology, morphology, and ocular age and rage.

PURPOSE: To determine the effects of the advanced glycation end product (AGE) cross-link breaker alagebrium on intraocular pressure (IOP), accommodation (ACC), outflow facility (OF), anterior segment morphology, and ocular AGE and receptors for AGE (RAGE) in older rhesus monkeys. METHODS: Six rhesus monkeys (aged 19 to 20 years) received 3 or 4 intracameral and intravitreal (final concentration, 1 mM) injections of alagebrium to one eye over 2.5 to 3 weeks and vehicle to the opposite eye. ACC and OF responses to intramuscular or intravenous pilocarpine were measured at baseline and at 1 to 2 weeks and 2, 4, and 6 months postinjection. IOP was measured prior to all injections, ACC, and OF measurements. Monkeys were euthanized 3 to 6 months after the last injection, the eyes were enucleated, and anterior and posterior segments were examined by electron microscopy or immunohistochemistry. RESULTS: No significant differences were found in ACC or IOP at any time point after alagebrium treatment. Baseline OF was higher (37.0 +/- 6.0%; P < or = .005) in alagebrium-treated vs control eyes at 6 months postinjection. In 3 monkeys, alagebrium-treated eyes, compared to control eyes, showed greater focal plaque formation, similar to that seen in primary open-angle glaucoma, in the juxtacanalicular meshwork/inner wall of Schlemm's canal. No changes in anterior segment AGE or RAGE were detectable. However, some areas of the retina and optic nerve head exhibited decreased AGE and increased RAGE immunostaining. CONCLUSIONS: Intraocular injection of AGE cross-link breakers is an unlikely approach for glaucoma therapy. However, it may generate a model for further study of glaucomatous-like plaque formation. Immunohistochemical changes in the posterior segment in response to alagebrium warrant further functional studies.

Identification of unknown intraocular material after cataract surgery: evaluation of a potential cause of toxic anterior segment syndrome.

PURPOSE: To describe and identify unknown opaque material between the optic of an AR40 intraocular lens (IOL) injected with the Emerald Series implantation system (both AMO, Inc.) and the posterior capsule at the conclusion of routine phacoemulsification to prevent an outbreak of toxic anterior segment syndrome (TASS). SETTING: Ambulatory care center operating room, University of North Carolina Hospitals and Department of Ophthalmology, University of North Carolina School of Medicine at Chapel Hill, Chapel Hill, North Carolina, USA. METHODS: After coaxial phacoemulsification in multiple patients, opaque material was present between the optic of a posterior chamber IOL and the posterior capsule. Although there was no TASS, the material was removed from 2 eyes and analyzed with scanning electron microscopy (SEM) and x-ray microanalysis (XRM). Similarly, crystalline lens, Klenzyme (Steris Corp.), Viscoat (sodium hyaluronate 3.0%-chondroitin sulfate 4.0%), and Provisc (sodium hyaluronate 1.0%) were analyzed. RESULTS: On SEM, the material had an irregular undulating surface similar to that of Provisc. Viscoat and the crystalline lens had smoother surfaces. On XRM, the material contained sodium, chlorine, and calcium, like Viscoat and Provisc, and phosphorous and sulfur, like Viscoat. The material also contained silicone, magnesium, aluminum, titanium, iron, and zinc. Klenzyme had smaller peaks of sodium, chlorine, and calcium and a higher carbon background than the unknown material. CONCLUSIONS: The material was likely ophthalmic viscosurgical device that was chemically and structurally altered by the cleaning and sterilization process. The silicone and metallic elements were probably from the Emerald Series implantation system as the disposable cartridge is coated with silicone and the reusable injector is metal.

Canaloplasty: circumferential viscodilation and tensioning of Schlemm's canal using a flexible microcatheter for the treatment of open-angle glaucoma in adults: interim clinical study analysis.

PURPOSE: To evaluate the safety and efficacy of circumferential viscodilation and tensioning of the inner wall of Schlemm's canal in a new surgical procedure for the treatment of open-angle glaucoma (OAG). SETTING: Fourteen clinical sites in the United States and Germany. METHODS: In this international multicenter prospective study of adult patients with OAG having glaucoma surgery, patients with qualifying preoperative intraocular pressure (IOP) of at least 16 mm Hg or higher and open angles were eligible. Evaluation was performed at baseline and 1 day, 1 week, and 1, 3, 6, and 12 months preoperatively. After a nonpenetrating dissection technique to expose Schlemm's canal was performed, a flexible microcatheter (iTrack 250A, iScience Interventional) was used to dilate the full circumference of the canal by injecting sodium hyaluronate 1.4% (Healon GV) during catheterization. A suture loop was placed in the canal to apply tension to the trabecular meshwork. High-resolution ultrasound imaging was used to assess Schlemm's canal and anterior segment angle morphology, including distension of the trabecular meshwork caused by the tensioning suture. Data analysis was performed in 2 groups: Group 1, in which patients met all inclusion criteria, and Group 2, made up of Group 1 patients who had successful suture placement. RESULTS: Group 1 comprised 94 patients and Group 2, 74 patients. The mean baseline IOP in Group 1 was 24.7 mm Hg+/-4.8 (SD) on a mean of 1.9+/-1.0 medications per patient. In Group 2 (patients with sutures), the mean IOP was 16.1+/-4.7 mm Hg 3 months postoperatively, 15.6+/-4.0 mm Hg at 6 months, and 15.3+/-3.8 mm Hg at 1 year. Medication use dropped to a mean of 0.6+/-0.9 per patient at 12 months. Suture tensioning was an apparent contributing factor in achieving surgical success. Patients with measurable trabecular meshwork distension from suture tension had a mean IOP of 15.9+/-5.2 mm Hg at 6 months and 14.5+/-3.0 mm Hg at 12 months. Surgical and postsurgical adverse events were reported in 15 of 94 patients (16%) and included hyphema (3), elevated IOP greater than 30 mm Hg (3), Descemet's tear (1), hypotony (1), choroidal effusion (1), and exposed closure suture with eyelid edema and erythema epiphora (1); 4 patients were subsequently converted to trabeculectomy. CONCLUSION: Circumferential viscodilation and tensioning of Schlemm's canal was a safe and effective surgical procedure to reduce IOP in adult patients with OAG.

Col4a1 mutation causes endoplasmic reticulum stress and genetically modifiable ocular dysgenesis.

Ocular anterior segment dysgenesis (ASD) is a complex and poorly understood group of conditions. A large proportion of individuals with ASD develop glaucoma, a leading cause of blindness resulting from retinal ganglion cell death. Optic nerve hypoplasia is thought to have distinct causes and is a leading cause of blindness in children. Here, we show that a mutation in the type IV collagen alpha 1 (Col4a1) gene can cause both ASD and optic nerve hypoplasia. COL4A1 is a major component of almost all basement membranes. The mutation results in non-secretion of the mutant COL4A1 proteins, which instead accumulate within cells. Basement membrane abnormalities may, therefore, contribute to the phenotype. The mutation also induces endoplasmic reticulum stress and so intracellular stress may contribute to pathogenesis. The overall consequence of the Col4a1 mutation depends on genetic context. In one genetic context, the mutation causes severe ASD with intraocular pressure abnormalities and optic nerve hypoplasia. In a different genetic context, both the ASD and optic nerve hypoplasia are rescued, and we have identified a single dominant locus that confers the phenotypic modification.

Exploración del segmento anterior mediante tomografía óptica de coherencia 3 / Exploration of the anterior segment by optical coherence tomography 3

Objetivo: Dar a conocer la utilidad de la Tomografía de Coherencia Óptica (OCT) para el estudio de patología del segmento anterior así como presentar sus posibles ventajas frente a la biomicroscopía ultrasónica (BMU). Material y métodos: Presentamos 5 pacientes con diversas patologías del segmento anterior estudiados mediante OCT: 2 pacientes con ataque agudo de glaucoma de ángulo estrecho, 1 paciente con rubeosis iridis, 1 paciente con un traumatismo ocular penetrante y un paciente con un quiste estromal primario del iris. Resultados: En los pacientes con ataque agudo de glaucoma, el OCT ha mostrado ser una herramienta tan útil como la BMU para la detección del cierre angular. Además, la tomografía óptica fue más cómoda y rápida en la obtención de las imágenes que la BMU, así la el tiempo medio de exploración con OCT suele ser inferior a 5 minutos, a diferencia de la BMU cuyo tiempo suele ser superior a 10 minutos. El OCT también ha demostrado ser una prueba útil para el estudio de otras patologías del iris, permitiendo valorar la permeabilidad de las iridotomías, mostrar neovascularización iridiana e incluso estudiar masas iridianas como el quiste estromal de iris. En estos casos, el OCT permite la obtención de imágenes con una mayor resolución espacial que la BMU. Conclusiones: El OCT 3 mediante el ajuste del enfoque puede servir para obtener imágenes del ángulo camerular así como de otras estructuras del segmento anterior ayudando así a establecer determinados diagnósticos de forma no invasiva, cómoda y segura para el paciente

Purpose: To show the utility of optical coherence tomography (OCT) for studying the anterior segment and to explain its potential advantages as compared to ultrasonic biomicroscopy (UBM). Materials and methods: We have described the findings in 5 patients with different pathologies of the anterior segment, all of whom were studied with UBM and OCT-3 adjusted for the anterior segment. There were 2 cases of an acute attack of closed angle glaucoma, 1 case of rubeosis iridis, 1 case of penetrating ocular trauma and 1 case of a primary stromal iris cyst. Results: OCT was found to be as useful as UBM in detecting angle closure in patients affected by an acute attack of glaucoma. In addition, OCT was more comfortable for the patient and faster than UBM in obtaining images, with the exploration mean time using OCT being less than five minutes while that with UBM was over ten minutes. OCT has also been demonstrated to be a safe and valuable non-contact examination in other iris pathologies such as rubeosis iridis, for checking the permeability of iridotomies and even for studying iris masses such as primary stromal iris cysts. In these entities OCT may reach a higher resolution than ultrasonic biomicroscopy. Conclusion: OCT-3, with an adjustable focus, can obtain images from the scleral angle, as well as from other ocular structures like the iris, thus assisting in the diagnosis of numerous pathologies

Role of ultrasound biomicroscopy (UBM) in the detection and localisation of anterior segment foreign bodies.

INTRODUCTION: The aim of this study was to investigate the role of ultrasound biomicroscopy (UBM) in imaging anterior segment foreign bodies and compare it with conventional B-scan ultrasound and computed tomography (CT). MATERIALS AND METHODS: The charts of 18 eyes with anterior segment foreign bodies were reviewed. The rates of detection of foreign bodies using ultrasound, CT scan and UBM were compared. RESULTS: The foreign body detection rates were 36.5% by ultrasound, 88.9% by CT scan, and 94.4% by UBM. The diagnosis of presence of a foreign body using UBM was made based on high reflective echoes causing shadowing or reverberations. In 7 eyes, UBM detected injury to the zonules, guiding the operative procedure. In the 8 eyes for which all tests were performed, rates of detection of foreign bodies were 25% (2/8) with ultrasound, 87.5% (7/8) with CT, and 100% (8/8) with UBM. CONCLUSION: UBM is a valuable adjunct for the accurate localisation of small foreign bodies, including cilia. It offers a higher detection rate than that provided by ultrasound and CT scan.

Vehicle of triamcinolone acetonide is associated with retinal toxicity and transient increase of lens density.

BACKGROUND: Intravitreal triamcinolone acetonide (TA) has been widely used as a therapeutic method for many ocular diseases, but a consensus on an appropriate safe therapeutic window of dosage for intravitreal injection, and whether vehicle of TA should be reduced or eliminated, has not yet been reached. The aim of this article is to investigate these issues. METHODS: Forty New Zealand white rabbits were divided into four experimental groups and one control group. Four or 25 mg TA, with vehicle either reduced or not, was injected into the vitreous cavity of rabbits in experimental groups. Rabbits in the control group received 0.2 ml intravitreal sterile saline solution. Intraocular pressures (IOP) were measured by a Tonopen tonometer. Values of lens density were measured by a Pentacam system. Soluble protein, total antioxidation capacity, reduced glutathione (GSH), glutathion peroxidase (GSH-px), and superoxide dismutase (SOD) in lens were measured by specific kits. ERG and pathological examinations, including light and electron microscopy of the retina, were also performed. RESULTS: Elevation of IOP was noted in all experimental groups after intravitreal TA (P<0.01, paired t-test). Significant increase of lens density was noticed at 1 week after intravitreal TA in the 25 mg vehicle-containing group (P<0.0001, paired t-test). Significant loss of GSH-px activity was noticed at the end of the study (P<0.05, paired t-test), while SOD activity increased (P<0.05, paired t-test). Amplitudes of ERG waves declined significantly in vehicle-containing groups (P<0.01, paired t-test) at the end of the study. Pathological examination showed obvious retinal toxicity in vehicle-containing groups. CONCLUSIONS: Vehicle of TA should be eliminated or reduced before intravitreal injection to avoid potential retinal toxicity and transient increase in lens density.

Do injections of 5-fluorouracil after trabeculectomy have toxic effects on the anterior segment?

OBJECTIVE: To discourage fibrosis of the filtering bleb, 5 fluorouracil (5-FU) may be injected after trabeculectomy. 5-FU is an antimetabolite that also can damage extraocular tissues at concentrations as low as 0.5%. This study ascertained whether repeated injection of 5-FU has toxic effects on intraocular structures. METHODS: After unilateral trabeculectomy in anesthetized New Zealand rabbits, 5-FU (5.0 mg/0.1 mL) was injected at the trabeculectomy site every 5 days for 15 days. Evaluation included slit-lamp examination, confocal microscopy, and intraocular pressure (IOP). After sacrifice, aqueous humor (AH) was drawn and eyes excised for scanning electron microscopy (SEM) and light microscopy. RESULTS: The 5-FU injection not decrease IOP beyond trabeculectomy alone. Bleb height remained constant, thickness increased, and vascularity decreased. No changes in cornea or anterior segment were observed. No inflammation was observed in the bleb or surrounding tissues by slit-lamp or histologic examination. Protein in AH increased from 0.6 +/- 0.5 microg/mL at baseline to 19.8 +/- 4.4 microg/mL after trabeculectomy but only to 0.9 +/- 0.6 microg/mL after trabeculectomy plus 5-FU. Both in vivo confocal microscopy and SEM revealed deleterious effects on corneal epithelial and endothelial cells with a minor shift toward smaller cells. CONCLUSIONS: In this study 5-FU did not provoke an intraocular inflammatory response and had minimal effect on extraocular structures. Changes in corneal epithelium and endothelium detectable by confocal microscopy suggest a small toxic effect. These in vivo measurements by confocal microscopy were confirmed by SEM. Repeated administration did not cause additional cumulative toxic effects in the anterior segment. Therefore, multiple injections of 5- FU into the filtering bleb pose minimal risk to intraocular structures.

[Intraobserver reproducibility in anterior segment morphometry of normal eyes using ultrasound biomicroscopy (UBM)]. / Reprodutibilidade intra-observador de medidas morfométricas do segmento anterior em olhos normais pela biomicroscopia ultra-sônica (UBM).

PURPOSE: To systematically study the anterior segment morphometric measurements by ultrasound biomicroscopy (UBM) in a large population of normal eyes, to evaluate their intra-observer reproducibility. Two new measurements are proposed and also tested concerning their intra-observer reproducibility. METHODS: One hundred ninety eyes of 101 patients with normal ophthalmic examination were studied by UBM. Anterior chamber depth (ACD) along with 11 different morphometric parameters that characterize the anterior segment morphometry were measured in the superior, nasal, inferior and temporal meridians. All measurements were performed twice with a minimum interval of four weeks between them. RESULTS: There were no statistical significant differences between the first (M1) and second (M2) measurements and regarding each studied parameter except for 2 variables in 2 meridians in the right eyes (OD) and 2 variables of one meridian in the left eyes (OS). Even in these cases, the largest difference between M1 and M2 was less than 0.006 mm, which was not clinically significant. The new tested parameters also presented a good intra-observer reproducibility. CONCLUSION: This study confirmed the good intra-observer reproducibility of the variables that characterize anterior segment morphometry by UBM.

Reprodutibilidade intra-observador de medidas morfométricas do segmento anterior em olhos normais pela biomicroscopia ultra-sônica (UBM) / Intraobserver reproducibility in anterior segment morphometry of normal eyes using ultrasound biomicroscopy (UBM)

OBJETIVO: Estudar as variáveis que caracterizam a morfometria do segmento anterior usando a biomicroscopia ultra-sônica (UBM), em pacientes com o exame oftalmológico normal, em amostra de tamanho significativo, de forma sistematizada, com o intuito de avaliar sua reprodutibilidade intra-observador. Dois novos parâmetros tiveram sua reprodutibilidade intra-observador também testada. MÉTODOS: Foram examinados 190 olhos de 101 pacientes com exame oftalmológico normal empregando-se a UBM. Em cada olho, além da profundidade da câmara anterior (PCA), onze outros parâmetros que caracterizam a morfometria do segmento anterior foram medidos nos meridianos superior, nasal, inferior e temporal, em dois momentos distintos com intervalo mínimo de quatro semanas entre eles, pelo mesmo observador. RESULTADOS: Não se observaram diferenças estatisticamente significativas entre a primeira e a segunda medida (M1 e M2) de cada parâmetro estudado, exceto quanto a duas variáveis em dois meridianos nos olhos direitos (OD), e duas variáveis em um meridiano nos olhos esquerdos (OE). Mesmo estas diferenças mostraram-se clinicamente não significantes, por serem inferiores a 0,006 mm. Os dois novos parâmetros testados também apresentaram boa reprodutibilidade intra-observador. CONCLUSAO: Este estudo confirmou a boa reprodutibilidade intra-observador das variáveis que caracterizam a morfometria do segmento anterior pela UBM.

Factors influencing intraocular pressure in cultured human anterior segments.

PURPOSE: To determine why variations in intraocular pressure (IOP) occur in cultured human anterior segments despite a constant rate of infusion of culture medium. Two types of variations occur: an initial elevation of IOP and small changes in baseline IOP. METHODS: Anterior segments from human eyes were placed in perfusion organ culture. In cultures with initially high IOP, eyes were fixed at the high IOP level and histologic examination performed. In other cultures with high initial IOP, effluent medium was collected and subsequently reinfused after IOP had decreased to baseline. In cultures with stable baseline IOP, cell fragments from monolayer-cultured cells, or human genomic DNA, were infused at concentrations equivalent to 30,000 to 300,000 cells. RESULTS: Electron microscopy of initially high-pressure cultures revealed scattered cell debris throughout the meshwork in greater amounts than found in eyes without initially high IOP. Reinfusion of effluent media from cultures with high initial pressures caused elevation of IOP. Centrifugation of effluent media lessened this elevation of IOP. In cultures with stable baseline IOP, infusion of cell fragments or genomic DNA raised IOP in a dose-dependent manner, with elevation of IOP for a minimum of 24 hours. CONCLUSIONS: Cell debris can elevate IOP during the initial culture period, and after baseline pressures are established. Cell fragments and DNA increase IOP in a dose-dependent manner. The variations in baseline IOP seen during culture are probably caused by cell fragments and debris from dying cells in the meshwork, ciliary body, and other anterior segment tissues.

Secondary glaucoma and severe endothelial damage after silicone phakic posterior chamber intraocular lens implantation.

We report a patient who developed secondary glaucoma 1 year after phakic posterior chamber intraocular lens implantation.

Histopathological study of the anterior lens capsule with a double-ring sign.

BACKGROUND: A 'double-ring' sign may be seen during continuous curvilinear anterior capsulorhexis in cataract surgery. This sign has been attributed to partial splitting of the anterior lens capsule at the continuous curvilinear anterior capsulorhexis edge. Because horizontal intracapsular splits away from the capsule edge have also been reported, a double-ring sign has been hypothesized to be a precursor to true exfoliation. We report new histopathological findings for both anterior capsule and lens epithelium changes, the latter being similar to those of true exfoliation. METHODS: Three anterior capsules from two patients with a double-ring sign during capsulorhexis were examined via light and transmission electron microscopy. RESULTS: Light microscopy revealed step formations in the capsule edges and surface-parallel intracapsular splits in the anterior capsules. Transmission electron microscopy revealed stratified, banded structures consisting of alternating electron-dense and -lucent granular belts (300-400 nm) at the central area of the anterior capsule, as well as vacuole-like spaces and expansion of intercellular spaces in the lens epithelium. CONCLUSIONS: The thickened, stratified structure of the anterior capsule suggests abnormal lens epithelium in patients with a double-ring sign. The horizontal splits in the anterior capsule, with concomitant epithelial changes, i.e., the vacuole-like spaces and widened intercellular spaces, support the relation between a double-ring sign and true exfoliation.

Effect of heparin II domain of fibronectin on aqueous outflow in cultured anterior segments of human eyes.

PURPOSE: To determine whether an integrin/syndecan-binding domain of fibronectin, called the heparin II (Hep II) domain, affects outflow facility in the human eye. METHODS: Anterior segments of human eyes were placed in perfusion organ culture. One eye of each pair received the Hep II domain, and the fellow eye received DMEM or a heat-denatured Hep II domain. The Hep II domain was produced as a recombinant glutathione S-transferase (GST)-fusion protein. Microscopic changes were assessed. RESULTS: Outflow facility in anterior segments treated with Hep II domain increased by 93% compared with that in anterior segments treated with DMEM. In contrast, facility in anterior segments treated with the heat-denatured Hep II domain showed very little change. Outflow facility remained high during Hep II domain perfusion and returned to baseline after removal of the protein. Electron microscopy revealed disruptions in the endothelial lining of Schlemm's canal in anterior segments fixed during maximum effect and in anterior segments after facility had returned to baseline. Scattered disruptions of canal cells were noted in control anterior segments. Trabecular cells in other regions looked normal. Major changes in the extracellular matrix of the juxtacanalicular tissue were not observed. Repeated doses of the Hep II domain administered after facility returned to baseline increased facility in two of three anterior segments. CONCLUSIONS: The Hep II domain of fibronectin increases outflow facility in the human anterior segment. This suggests that fibronectin-mediated interactions may have a role in modulating aqueous hydrodynamics. Such interactions may represent avenues of novel therapeutic interventions for glaucoma.

The mechanism of increasing outflow facility during washout in the bovine eye.

PURPOSE: To investigate the relationship between outflow facility and separation between the inner wall of the aqueous plexus and the juxtacanalicular connective tissue (JCT) during washout in the bovine eye. METHODS: Facility was recorded during 3 hours of anterior chamber perfusion at 15 mm Hg in eight pairs of bovine eyes. One eye of each pair was then lowered to 0 mm Hg for 1 hour, whereas the fellow eye was kept at 15 mm Hg. After a brief perfusion at 15 mm Hg, both eyes were perfusion fixed and processed for electron microscopy. Micrographs of the inner wall were analyzed for separation from the JCT. To study the role of cellular adhesion between the inner wall and JCT, 12 additional pairs were perfused with integrin-binding peptide (RGD: Arg-Gly-Asp) or sham control peptide (RGE: Arg-Gly-Glu) at 2 micro M to 2 mM, before IOP was reduced. RESULTS: During the first 3 hours, facility increased in both eyes because of "washout." However, after 1 hour of 0 mm Hg, facility decreased by 13% (P < 0.006), whereas facility increased by 20% (P < 0.001) in the fellow eyes maintained at 15 mm Hg. Two types of separation were observed between the inner wall and JCT: cell-matrix separation between the endothelial cell and basal lamina and matrix-matrix separation between the basal lamina and JCT. A significant positive correlation (P = 0.042) was found between the degree of matrix-matrix separation and the change in outflow facility after 1 hour of 0 mm Hg. Compared with RGE control, RGD had no apparent effect on outflow facility (P > 0.35) or on the change in outflow facility after 1 hour at 0 mm Hg (P > 0.15). CONCLUSIONS: The increase in outflow facility that occurs during washout in the bovine eye is reversible and correlates with the degree of separation between the basal lamina of the inner wall endothelium and the JCT. Therefore, adhesions tethering the inner wall to the JCT may be important ultrastructural features involved in the regulation of aqueous humor outflow resistance.