Localization of transport adenosine triphosphatase in rat cornea.

Increasingly, physiological and biochemical data on the mammalian cornea have emphasized the importance of the endothelial rather than the epithelial layer in regulating the state of hydration of the cornea, via a transport ATPase. This view is supported by the observations reported here. With the Ernst method, the first cytochemical procedure to be readily responsive to ions and ouabain, the major sites of enzyme reaction product are the intercellular spaces of the endothelial layer. This localization was not found by us with the so-called ATPase method of Wachstein and Meisel.

Studies on microperoxisomes. VII. Pigment epithelial cells and other cell types in the retina of rodents.

The pigment epithelial cell of the retina actively participates in two aspects of lipid metabolism: (a) the fatty acid esterification of vitamin A and its storage and transport to the photoreceptors, and (b) the phagocytosis and degradation of the lipoprotein membrane disks shed from the photoreceptor cells. Study of the pigment epithelial cells of adult albino and pigmented rodents has revealed the abundance of an organelle, microperoxisomes, not previously known to exist in this cell type. The metabolism, transport, and storage of lipids are major functions of other cell types which possess large numbers of microperoxisomes associated with a highly developed smooth endoplasmic reticulum. Microperoxisomes were encountered, but relatively rarely, in Müller cells and vascular endothelial cells. A tubular system in photoreceptor terminals is reactive in the cytochemical procedure used to visualize microperoxisomes.

Exocytic clearing of degraded membrane material from pigment epithelial cells in frog retina.

Posterior eye cups of the frog Rana ridibunda were prefixed in the presence of tannic acid, osmicated and stained en bloc with uranyl acetate. Electron microscopy revealed the following features of membrane traffic in cells of the retinal pigment epithelium (RPE): (1) intracellular membranes with increased electron density undergo degradation within phagolysosomes and also outside lysosomes and are committed to be exocytized; (2) exocytized degraded membranes, organized as lamellar arrays, are evacuated through the basolateral extracellular space, through Bruch's membrane and into the choriocapillaris. Comparison of exocytic activity in animals kept either with a light cycle or in constant darkness suggests that exocytized lamellar arrays derive from membranes degraded through both lysosomal and non-lysosomal pathways. Autophagocytosis may contribute a substantial amount of material to exocytosis. On the other hand, exocytosis of heterophagosomes, if it takes place at all, involves only residual bodies at a final stage of degradation following slow proteolysis which is achieved within the RPE cells. Rapid transcytosis of heterophagosomes does not occur. Taken together, our observations suggest that, in the frog, exocytosis and subsequent evacuation of residual bodies is an efficient mechanism for clearing from worn-out organelles the metabolically highly active, non-dividing RPE cells.

Effect of laser photocoagulation on oxygenation of the retina in miniature pigs.

A large area of the posterior pole of the retina of the miniature pig was photocoagulated, and 2-3 wk later the PO2 in the preretinal vitreous was mapped with O2-sensitive microelectrodes. In the control retina, the PO2 was highly heterogeneous being much higher close to an artery than opposite an intervascular zone. After photocoagulation, PO2 opposite an intervascular zone was found to be significantly increased. A quantitative histologic analysis showed that in the photocoagulated areas more than 28% of the outer retina was destroyed. The authors conclude from these results that photocoagulation, by partially destroying the pigment epithelium-photoreceptor complex, causes an increase of the oxygenation of the retina.

Glial reactivity, an early feature of diabetic retinopathy.

PURPOSE: To characterize early structural gliotic reactions in retinal Müller cells, astrocytes, and microglia in experimentally induced diabetes. METHODS: Rats were rendered diabetic by streptozotocin injection and killed after 2, 4, 12, or 20 weeks. Cell densities were determined in flatmounted retinas or transverse semithin sections. Expression of glial fibrillary acidic protein (GFAP) was localized on frozen sections or flatmounts by immunofluorescence and confocal microscopy, and GFAP content was evaluated by Western blot analysis. Microglial cells were visualized by binding of isolectin B4 or staining with antibodies to phosphotyrosine residues. The integrity of the blood-retinal barrier was assessed by intravenous injection of Evans blue. RESULTS: The density of Müller cells and microglia was significantly increased at 4 weeks of diabetes compared with nondiabetic controls. GFAP expression in Müller cells was not detected at 4 weeks but was prominent at 12 weeks. The number of astrocytes was significantly reduced at 4 weeks in the peripapillary and far peripheral retina. Shape changes of microglial cells indicated functional activation. Leakage of the blood-retinal barrier was observed at 2 weeks of hyperglycemia, the earliest time point investigated. CONCLUSIONS: The leakage of the blood-retinal barrier before glial reactivity suggests that glia are early targets of vascular hyperpermeability. The individual glial cell types react differentially to the diabetic state. Müller cells undergo hyperplasia preceding GFAP expression, and microglial cells are activated, whereas astrocytes regress. This glial behavior may contribute decisively to the onset and development of neuropathy in the diabetic retina.

Remodeling of retinal capillaries in the diabetic hypertensive rat.

PURPOSE: To document the effect of sustained systemic hypertension on the integrity and ultrastructural morphology of retinal capillaries in diabetic and nondiabetic rats. METHODS: Normotensive (strain Wistar-Kyoto; WKY) and genetically hypertensive (spontaneously hypertensive; SHR) rats were rendered diabetic by intravenous streptozotocin injection. At 20 weeks of diabetes, endothelial cells, pericytes, and extracellular matrix were evaluated by ultrastructural morphometry. Serum albumin was localized by immunofluorescence microscopy. RESULTS: The endothelial cell layer was markedly thinner in the diabetic normotensive animals. The number of intercellular junctions was reduced in both the nondiabetic and diabetic hypertensive group but less so in the diabetic normotensive group. No significant endothelial cell loss was noted in either of the experimental groups, whereas the number of pericytes and the number of their cytoplasmic processes were reduced in diabetic and hypertensive animals. Significant thickening of the basement membrane and increased permeability to serum albumin were observed in diabetic and hypertensive rats and were strongly enhanced in the combined diseases. CONCLUSIONS: Endothelial thinning and shape changes from an elaborate to a simpler form as well as rounding up of the pericytes and loosening of their vascular sheaths indicate remodeling of the vascular wall during chronic diabetes and sustained hypertension, before a characteristic vasculopathy becomes manifest. The combination of diabetes and hypertension enhances these features, as well as basement membrane thickening and breakdown of the blood-retinal barrier.

Expression and release of tumor necrosis factor-alpha by explants of mouse cornea.

PURPOSE: To elucidate a possible target of immunosuppressive agents widely used in the treatment of corneal disorders, the authors determined whether corneal cells are capable of expressing and releasing tumor necrosis factor-alpha (TNF alpha) on lipopolysaccharide (LPS) stimulation, and they investigated whether TNF alpha production can be modulated by pharmacologic agents. METHODS: Trephined central corneas from C57BL/6 mice were kept in culture for 3 days. Release of TNF alpha after a 24-hour stimulation with LPS (1 microgram/ml) into the culture medium was determined both by bioassay and by enzyme-linked immunosorbent assay. Expression of TNF alpha mRNA after 6-hour stimulation was examined by polymerase chain reaction. Immunofluorescent staining on cryostat sections of cultured corneas was performed to localize TNF alpha in the tissue. Corneal explants were pretreated with immunosuppressive agents (prednisolone, budesonide, cyclosporin A) for 48 hours, followed by 6-or 24-hour stimulation with LPS in the continuous presence of the agents. RESULTS: Lipopolysaccharide stimulated TNF alpha release into the culture medium. The addition of budesonide (10(-7) M) or prednisolone (10(-6) M) significantly inhibited LPS-induced TNF alpha release, whereas cyclosporin A (10(-7) - 10(-5) M) had no marked effect. Levels of TNF alpha mRNA in corneal explants increased fivefold after stimulation with LPS. Immunohistochemical staining revealed that TNF alpha was expressed in the epithelial cells. Budesonide markedly decreased mRNA expression and abolished immunostaining of TNF alpha stimulated by LPS. CONCLUSIONS: TNF alpha is produced and released by the epithelial cells of mouse central cornea in response to LPS. Contrary to cyclosporin A, corticosteroids such as prednisolone and budesonide potently inhibit TNF alpha production.

Contrast sensitivity in obese dyslipidemic patients with insulin resistance.

OBJECTIVE: To evaluate contrast sensitivity in insulin-resistant obese patients and in aretinopathic diabetic patients. METHODS: Contrast sensitivity was measured at 3 letter sizes (44 x 44, 9 x 9, and 5 x 5 mm) in mesopic (5 candela [cd]/m2) and low photopic (85 cd/m2) vision in 20 dyslipidemic obese patients with insulin resistance, 20 age-matched patients with type 2 diabetes mellitus, and 20 aged-matched healthy control subjects. RESULTS: Significant loss of contrast sensitivity at all 3 letter sizes was observed in low photopic vision (at 44 x 44-mm letter size, control vs obese, P<.002, and control vs diabetic, P<.005; at 9 x 9-mm letter size, control vs obese, P<.02, and control vs diabetic, P<.001; and at 5 x 5-mm letter size, control vs obese, P<.05, and control vs diabetic, P<.005) and mesopic vision (at 44 x 44-mm letter size, control vs diabetic, P<.005; at 9 x 9-mm letter size, control vs obese, P<.005, and control vs diabetic, P<.01; and at 5 x 5-mm letter size, control vs obese, P<.005, and control vs diabetic, P<.001) in insulin-resistant obese and diabetic patients. CONCLUSION: The results suggest that an early neurosensory dysfunction may occur without visible vascular involvement and without overt hyperglycemia.

In vivo effects of glycine on retinal ultrastructure and averaged electroretinogram.

Glycine, when injected intravitreally, has an inhibitory action on the electroretinogram (ERG) of the rabbit resulting in a transient loss of the oscillatory potentials of the b-wave. This inhibitory action is reversible within 24 h after glycine injection. In autoradiographs, after [3H] glycine administration, the radioactive label is mainly found over the inner nuclear layer (INL) and inner plexiform layer (IPL) without predilection of specific cell types and/or synapses. Electron microscopy reveals cytopathological changes in amacrine cells, in particular their cell membranes. These changes are conspicuous especially 1-2 h after the injection and the cells become normal again within 24 h. It is concluded that glycine has an inhibitory action upon the rabbit ERG in vivo. This action, on the basis of our morphological observations under our experimental conditions, may be due to an overall somatic membrane action rather than to an action as inhibitory neurotransmitter at the synaptic level. The reversible cellular lesions of amacrine cells after glycine administration with a concomitant, transient loss of oscillatory potentials (OP) supports the hypothesis that the cellular origin of OP is situated in amacrine cells.

Results of viscocanalostomy for primary open-angle glaucoma.

PURPOSE: Viscocanalostomy is a nonperforating filtering surgical procedure that may avoid postoperative complications common with standard trabeculectomy. This study was conducted to determine the surgical outcome of this procedure after a postoperative observation period of 12 to 36 months. METHODS: Interventional consecutive case series. In a prospective study, a consecutive series of 67 eyes of 67 patients with chronic primary open-angle glaucoma underwent VCS. Excluded were patients with angle closure glaucoma, post-traumatic, uveitic, neovascular, or dysgenetic glaucoma, as well as patients who needed combined cataract-glaucoma procedures. The patients were examined postoperatively on the first day, first week, at 1 month, and then at 3-month intervals through 36 months. At each visit, best-corrected visual acuity, intraocular pressure (IOP), and the appearance of the surgical wound, anterior chamber, and indirect funduscopy were recorded. RESULTS: Complete success was defined as IOP less than or equal to 20 mm Hg and greater than or equal to 30% IOP reduction without medical or additional surgical treatment compared with the IOP from a preoperative level with maximum tolerated medical therapy; qualified success IOP less than or equal to 20 mm Hg with treatment or an IOP reduction less than 30% from preoperative level with maximum tolerated medical therapy; and qualified failure of an IOP greater than 20 mm Hg with glaucoma medication, but no optic nerve or visual field deterioration and complete failure as an eye requiring further glaucoma surgery or lost visual function. The overall success rate was 88% at 1 year, 90% at 2 years, and 88% at 3 years, with a complete success of 68% at 1 year, 60% at 2 and 59% at 3 years. Four eyes had a perforation of the Descemet membrane, three of those needed peripheral iridectomy; six eyes had a microperforation not needing a peripheral iridectomy. Five eyes presented hyphema. Eight eyes presented deterioration of visual function after surgery: three eyes because of corneal astigmatism, one eye because of cataract, and four presented glaucomatous deterioration of the visual field despite pressures under 20 mm Hg. Four eyes had progressive cataract formation judged as independent from surgery. CONCLUSION: VCS provides an overall success rate of 88% and a complete success of 59% 3 years after surgery. The major immediate complication is perforation of the Descemet membrane with a need for peripheral iridectomy. No serious long-term complications were noted in our small series. Visual function remained stable in 55 eyes (82%).

Aqueous humor penetration of ofloxacin given by various routes.

We studied the aqueous humor penetration of ofloxacin after topical, oral, and intravenous administration in 51 consecutive patients undergoing cataract surgery. Aqueous humor concentration (mean +/- SD) was 0.53 +/- 0.35 mg/l when ofloxacin 0.3% eyedrops were instilled topically six times, one drop every three hours, until 90 minutes preoperatively, and 0.63 +/- 0.29 mg/l (P = .45) when two additional instillations were made, one drop every 30 minutes, until 30 minutes before aqueous humor aspiration. Aqueous humor concentration two hours after a single 200-mg oral dose (0.38 +/- 0.15 mg/l) was significantly lower (P = .048) than that 12 hours after the same oral dose (0.58 +/- 0.24 mg/l). Two hours following an intravenous infusion of 200 mg of ofloxacin, aqueous humor concentration was 0.33 +/- 0.19 mg/l. Our results suggest that therapeutic levels above the minimum inhibitory concentration for many bacteria cultured in endophthalmitis can be achieved in aqueous humor after either topical or oral administration, which indicates that this antibiotic passes easily through the corneal and the blood aqueous barriers.

Exfoliation syndrome and phacoemulsification.

PURPOSE: To evaluate intraoperative and postoperative complications of phacoemulsification and intraocular lens (IOL) implantation in eyes with exfoliation syndrome. SETTING: Eye Clinic, University Hospital of Geneva, Switzerland. METHODS: This prospective study evaluated 20 consecutive patients with exfoliation syndrome and 20 consecutive patients without the syndrome (control group) who had phacoemulsification and posterior chamber IOL implantation. One year later, the incidence of zonular tears, operating time, and fibrinoid reaction were studied in another group of 23 consecutive patients who had the same type of surgery by the same surgeon. RESULTS: Maximal pupil dilation was significantly smaller (P < .0001) in the exfoliation than in the control group. Intraoperatively, the incidence of zonular tears was the same in both groups (10%). In the 23 patients who had surgery 1 year later, no zonular tears occurred. The incidence of postoperative complications was similar in both groups. The exfoliation group had a significant decrease in intraocular pressure over preoperative levels at 1 week and 3 months postoperatively. CONCLUSION: Phacoemulsification with posterior chamber IOL implantation appears to be safe in eyes with exfoliation syndrome.

[Lamellar corneal graft in the treatment of pterygium. A 10-year retrospective study of the recurrence and changes of astigmatism]. / La greffe lamellaire cornéenne dans le traitement du ptérygion. Etude rétrospective sur 10 ans des taux de récidives et de la modification de l'astigmatisme.

57 eyes with a pterygium, including 12 eyes (22%) from which a pterygium had previously been removed by a simple excision, underwent a corneoscleral lamellar excision of pterygium followed by a corneal lamellar keratoplasty. During a mean follow-up of 37 months, we observed a recurrence in 2 eyes (3.5%). The corneal astigmatism measured with a Javal keratometer showed a modification in 54 eyes (95%). Visual acuity improved in 4 (7%) eyes, was unchanged in 29 (51%) and deteriorated in 23 (40%) eyes. The benefits of the low recurrence rate after this surgical procedure are to be weighed against the risk of modification of the corneal astigmatism and the possible modification of the visual acuity.

[Langerhans cells and corneal neovascularization in experimental alkali burns]. / Cellules de Langerhans et néovascularisation cornéenne dans les brûlures alcalines expérimentales.

Experimental alkali-burns rabbit corneas present a neovascularisation. Six weeks later the corneas have been processed for ATPase Cytochemistry, allowing the observation of Langerhans cells. We observed an increase in the number of Langerhans cells that was statistically significant in alkali-burned corneas when compared to unburned corneas in the peripheral and midperipheral cornea.

[Indications and techniques for removal of subluxated lenses (author's transl)]. / Cataracte subluxée, indications et techniques opératoires.

Extraction only appears indicated in cases of anteriour luxation and phakolytic glaucoma. Visual disturbances due to a dislocated lens should be considered relative indications for removal, since functional improvement cannot be expected as a rule. Expression of the lens through a corneoscleral incision is the method of choice in cases of anterior luxation (Fig. 1) as well as in subluxations without vitreous prolapse (Fig. 2) Open-sky vitrectomy allows for clearing of the route of extraction in cases of subluxation with vitreous in the anterior chamber (Fig. 3). A pars plana approach may be chosen in cases with a soft nucleus (Fig. 4) whereas luxated lenses with a hard nucleus are first aspirated (soft capsule and cortex), then the hard nucleus is luxated into the anterior chamber and finally extracted through a small corneoscleral incision, thus avoiding prolonged perfusion of the anterior chamber necessary for stripper techniques.