S-nitrosoglutathione photolysis as a novel therapy for antifibrosis in filtration surgery.

PURPOSE: To determine whether a novel peroxynitrite-based photosensitizer S-nitrosoglutathione (GSNO) can produce specific in vitro light-induced cell death of both standard animal lung and human Tenon's capsule (TC) fibroblasts and to compare this effect with that produced by the established photodynamic porphyrin precursor 5-aminolevulinic acid (ALA). METHODS: V79-4 Chinese hamster lung and human TC fibroblasts were established in tissue culture. GSNO, together with its radioactive tritiated and fluorescent dansylated derivatives, were synthesized. The labeled molecules were prepared to determine the time course of uptake into the fibroblasts. Uptake was monitored by scintillation counting for the tritiated GSNO and confocal fluorescence microscopy for the dansylated GSNO. The uptake of ALA and biosynthesis of its photosensitive product were determined by fluorescence emission spectroscopy of a separate set of fibroblasts. Once uptake was established, both cell lines were incubated with varying concentrations of GSNO or ALA as a function of time (0, 4, or 24 hours) before light exposure (200 msec pulsed visible light, 0.068 W per pulse, for 10 minutes at a distance of 10 cm). After 10 minutes of irradiation, the cells were washed and exposed to fresh tissue culture medium. The effect of the treatment was determined 24 hours later by measuring cell viability. RESULTS: A 2-minute drug treatment time (0 hours incubation) with GSNO, followed by 10 minutes of irradiation, resulted in approximately 78% of fibroblast cell death at the lowest concentration of GSNO used compared with the control, which was exposed to light, but no GSNO. The higher concentrations of GSNO, or longer drug treatment times before irradiation, did not statistically increase cell death. Maximal cell death was thus obtained using the lowest GSNO concentration (50 mM) and drug treatment time (2 minutes). In contrast, the well-established photosensitizer ALA killed only approximately 4% of cells at the lowest concentration and drug treatment time tested. At drug treatment times of 4 hours and less, increased concentrations of ALA did not produce cell death of more statistical significance. It was not until 24 hours of drug treatment that comparable amounts of cell death were produced by ALA and GSNO. In all experiments similar results were obtained with the animal lung and human TC fibroblasts, suggesting that the source of the fibroblast had no effect on the outcome. The differences in treatment effects between GSNO and ALA were statistically significant under all conditions tested. CONCLUSIONS: GSNO is able to cause light-specific cell death of human TC fibroblasts at drug treatment times (2 minutes) and irradiation times (10 minutes) that would be compatible with its use in glaucoma filtering surgery. This in vitro performance was superior to that of the well-established photosensitizer ALA, which required treatment times longer than 4 hours to approach the light-specific cell death produced by only 2 minutes of GSNO treatment.

Central corneal thickness in low-tension glaucoma.

BACKGROUND: It is possible that the intraocular pressure (IOP) is underestimated in eyes whose central cornea is thinner than normal. The objective of this study was to determine and establish the significance of central corneal thickness in patients with low-tension (normal-tension) glaucoma compared with those with chronic open-angle glaucoma (COAG) or ocular hypertension and healthy eyes. METHODS: The study was carried out from February 1998 to May 1999. Central corneal thickness was measured by ultrasonic pachymetry and IOP was measured by Goldmann applanation tonometry in 25 patients with low-tension glaucoma (untreated IOP less than 21 mm Hg with evidence of optic nerve head damage and corresponding visual field loss on automated perimetry), 80 patients with COAG (untreated IOP 21 mm Hg or greater with evidence of optic nerve head damage and corresponding visual field loss on automated perimetry), 16 patients with ocular hypertension (untreated IOP 21 mm Hg or greater, with normal optic nerve head and no history of glaucoma or elevated IOP, and normal visual field on automated perimetry) and 50 control subjects (untreated IOP less than 21 mm Hg with normal optic nerve head and no history of glaucoma or elevated IOP). Analysis with Pearson's product-moment correlation was performed to determine the correlation of IOP and central corneal thickness, and one-way analysis of variance was used to compare corneal thickness between groups. RESULTS: The central cornea was significantly thinner in the low-tension glaucoma group (mean 513.2 mu [standard deviation (SD) 26.1 mu]) than in the COAG group (mean 548.2 mu [SD 35.0 mu]) and the control group (mean 556.7 mu [SD 35.9 mu]) (p < 0.001). No significant difference in corneal thickness was found between the COAG and control groups. The ocular hypertension group had significantly thicker corneas (mean 597.5 mu [SD 23.6 mu]) than the three other groups (p < 0.001). INTERPRETATION: Patients with low-tension glaucoma may have thinner corneas than patients with COAG and healthy subjects. This results in underestimation of their IOP. Corneal thickness should be taken into account when managing these patients to avoid undertreatment.

Correlation of intraocular pressure and central corneal thickness in normal myopic eyes and after laser in situ keratomileusis.

PURPOSE: To determine the relationship of intraocular pressure (IOP) and central corneal thickness (CCT) in normal myopic eyes and after laser in situ keratomileusis (LASIK). SETTING: TLC The Windsor Laser Center, Windsor, Ontario, Canada. METHODS: Intraocular pressure measured by Goldmann applanation tonometry and CCT by ultrasonic pachymetry were determined in a group of untreated corneas of 120 patients (203 eyes) and in 50 patients (85 eyes) pre- and post-LASIK. Statistical analyses were performed with the Pearson correlation coefficient and paired Student t test. RESULTS: In the untreated group of 288 eyes, mean CCT was 544.0 microns +/- 37.3 (SD) (range 461 to 664 microns) and mean IOP, 15.6 +/- 2.7 mm Hg (range 10 to 24 mm Hg). The correlation between IOP and CCT in this group was highly significant (r = 0.44; P < .0001). The slope was 0.032 mm Hg/micron of CCT or an approximate decrease of 1 mm Hg, for a reduction in CCT of 31.3 microns. In the post-LASIK group, mean CCT dropped approximately 73.0 microns to 479.5 +/- 41.2 microns (range 408 to 503 microns) and IOP dropped to a mean of 13.6 +/- 3.3 mm Hg (range 7 to 22 mm Hg). A significant correlation was found between IOP and CCT after LASIK (r = 0.33; P < .002). The difference between the mean pre- and post-LASIK measurements of applanation IOP was 2.5 mm Hg, which was significant (P < .0001). The post-LASIK slope was 0.027 mm Hg/micron, or a decrease of 1.0 mm Hg per 37.8 microns reduction in CCT. CONCLUSION: Central corneal thickness is an important variable in the evaluation of applanation IOP and should be included in the assessment of any case of potential glaucoma or ocular hypertension, particularly in eyes with previous photoablative refractive surgery.

Stability of the antiproliferative effect of mitomycin-C after reconstitution.

PURPOSE: The purpose of this study was to determine the long-term stability of the antiproliferative effect of mitomycin-C (MMC) following reconstitution. METHODS: Identical MMC preparations were reconstituted from the crystalline form and then stored at room temperature or 4 degrees C. Cultured human Tenon's fibroblasts were exposed to MMC (0.5 mg/ml) for 2.5 min at 0, 3, 7, 10, 14, 21, 28, 35, and 42 days following reconstitution. After removal of the drug, fibroblast proliferation was measured by tritiated thymidine uptake. RESULTS: The percentage of inhibition was maintained at > 85% for both the room temperature and the 4 degrees C groups, and this inhibition persisted for the duration of the experiment. There was no statistically significant difference in the results between the storage temperatures. CONCLUSIONS: These data suggest that MMC retains its antiproliferative effect for at least 6 weeks following reconstitution and that this effect is not changed by storage temperature. Cost savings may be realized by storing MMC in unit-dose reconstituted aliquots for these longer periods, providing sterility can be assured.

Effects of cytoskeleton-reactive agents on aqueous outflow facility in a porcine ocular anterior segment preparation.

OBJECTIVE: To assess the effect of three agents known to react with cytoskeletal components--acrylamide (a vinyl monomer), demecolcine (a colchicine derivative) and stypoldione (an ichthyotoxic metabolite derived from the tropical marine alga Stypopodium zonale)--on facility of aqueous humour outflow in order to increase the understanding of the role of the cytoskeleton in the biology of aqueous outflow. DESIGN: Constant-pressure perfusion experiments with various concentrations of the agents using a porcine ocular anterior segment preparation. NUMBERS: Eighteen experimental eyes (six with each agent) and 18 control eyes. MEAN OUTCOME MEASURES: Aqueous outflow facility, determined after 1, 2 and 24 hours of perfusion. RESULTS: The mean baseline outflow facility was 0.10 (standard error of the mean 0.045) microL/min per mm Hg. There was no significant difference in mean baseline outflow facility between the experimental and control eyes. Perfusion of 0.01 to 1.0 mM demecolcine did not produce any consistent alteration in outflow facility. A mild to moderate dose-related experimental effect was noted after 2 hours of perfusion with 1.0 to 100 mM acrylamide. Perfusion with 0.10 mM stypoldione produced a moderate increase in outflow facility; the effect was greatest after 1 hour of perfusion. CONCLUSIONS: Our preliminary results do not support a central role for the cytoskeleton in the regulation of aqueous outflow resistance.

The effect of intracameral ethacrynic acid on the intraocular pressure of living monkeys.

Previous studies have shown that the sulfhydryl-reactive ethacrynic acid increases outflow facility in living monkeys when perfused via the anterior chamber. To study its potential clinical use further, living monkeys were intracamerally injected with 10 microliters of ethacrynic acid, with concentrations ranging from 0.5 to 7.5 mmol/l. The fellow control eye was injected with 10 microliters of diluent. The status of the anterior segment was monitored by slit-lamp biomicroscopy and the intraocular pressure was measured by pneumatonometry with the monkeys anesthetized with ketamine. The anterior segment of living monkeys tolerated injections up to 3.0-mmol/l ethacrynic acid without marked adverse effects. One of 13 monkey eyes injected with 3.0-mmol/l ethacrynic acid demonstrated mild reversible segmental corneal edema. The greatest mean intraocular pressure reduction in the 3.0- to 3.75-mmol/l group occurred at six hours, with the experimental intraocular pressure decreasing 2.9 mm Hg compared to a mean intraocular pressure increase of 0.1 mm Hg in the control group (n = 19). Concentrations of ethacrynic acid less than 3.0 mmol/l did not provide reliable reduction of intraocular pressure, whereas concentrations greater than 3.75 mmol/l caused a greater incidence and severity of corneal edema. We believe that the intracameral injection of ethacrynic acid can reliably and safely reduce intraocular pressure in living monkey eyes, and that this drug deserves further investigation as a potential antiglaucomatous agent.

Effects of topical ethacrynic acid adducts on intraocular pressure in rabbits and monkeys.

We evaluated the effect of topical ethacrynic acid on rabbit and monkey intraocular pressure. In a preliminary experiment, 100-mmol/L ethacrynic acid applied topically to Dutch-Belted rabbit eyes was associated with an 8-mm Hg lowering of intraocular pressure. However, corneal edema was severe, and the corneal epithelium sloughed off. To try to maintain the pressure-lowering effect but reduce the corneal side effects, we attempted to create an adduct of ethacrynic acid by utilizing ethacrynic acid's sulfhydryl reactivity. Ethacrynic acid was mixed with equimolar cysteine to bind the sulfhydryl-reactive sites on ethacrynic acid. The goal was to expose the cornea to adducted ethacrynic acid, which might then dissociate in the anterior chamber via a retro-Michael reaction. Intraocular pressure decreased 8.9 mm Hg (n = 40) with this treatment, and corneal edema was lessened (32 of 40 eyes had mild to no edema). However, we observed that when the eye was treated before ethacrynic acid-cysteine administration with topical acetylcysteine, the corneal side effects were reduced further and the intraocular pressure effect remained. In living cynomolgus monkeys receiving a single pretreatment drop of 75-mmol/L acetylcysteine followed by two drops of 130-mmol/L ethacrynic acid and 130-mmol/L cysteine, an intraocular pressure lowering of 9.9 mm Hg was observed (n = 7). However, in three of seven eyes corneal edema developed. Pretreatment with two drops of acetylcysteine eliminated the pressure-lowering effect but did not confer any added corneal protection. Our results indicate that topical ethacrynic acid-cysteine is effective in lowering the intraocular pressure of rabbits and cynomolgus monkeys and, when combined with acetylcysteine pretreatment, may offer the potential for a new topical therapeutic regimen for use in glaucoma.

Ocular sarcoidosis presenting as a solitary choroidal mass.

Ocular sarcoidosis presenting as a solitary choroidal mass without other signs of intraocular inflammation is a rare manifestation of systemic sarcoidosis. The authors describe a 26-year-old man who presented with a solitary choroidal tumour. Ocular and general physical examination as well as serum chemistry studies and pathological examination of a lymph node biopsy specimen confirmed sarcoidosis as the cause of the mass. Treatment with systemic steroids resulted in improvement in the appearance of the lesion; however, the visual acuity remained impaired. Other such cases reported in the literature are reviewed.

Corneal endothelial response to polymethylmethacrylate versus hydrogel lenses after phacoemulsification.

We performed a study to evaluate the corneal endothelial cell response in 26 patients who received a polymethylmethacrylate (PMMA) intraocular lens and 26 patients who received a soft, nonfolded poly-hydroxy-ethylmethacrylate (poly-HEMA) intraocular lens after posterior chamber phacoemulsification. Specular microscopy and pachymetry were done before surgery and a mean of 8 or more weeks after surgery. The mean percent cell loss was 8.2% in the PMMA group and 10.7% in the poly-HEMA group. There was no significant difference in the pachymetry values before or after surgery between the two groups; however, in the poly-HEMA group the postoperative value was significantly higher than the preoperative value (p = 0.027). The results suggest that the amount of perioperative corneal endothelial cell loss with poly-HEMA lenses is similar to that with PMMA lenses. Further study is needed to fully evaluate the long-term corneal effects of poly-HEMA lenses.