Previous studies have shown that sulfhydryl reagents can alter the facility of aqueous humor outflow but little is known about the sulfhydryl constituents of the aqueous outflow system or the effect of oxidants upon outflow facility. In the present study the concentration of glutathione (GSH) was measured in excised calf trabecular meshwork (TM) and found to be 0.40 mu mol/g wet wt (0.027 mu mol/mg protein). Oxidized glutathione was not detectable in the tissue. TM was found to have significant hexose monophosphate shunt activity as determined by measurement of the oxidation of 14C-1 and 14C-6-labeled glucose in tissue homogenates. The concentration of GSH in TM of enucleated calf eyes could be totally depleted by infusion of medium containing both diamide, which is an oxidant of GSH, and 1,3bis(2-chlorethyl)-1-nitrosourea (BCNU), which is an inhibitor of the enzyme glutathione reductase. The depletion of GSH was found to have no effect on the facility of aqueous outflow. Experiments were also done in which normal and TM GSH-depleted eyes were perfused with medium containing H202. Exposure to H202 produced no effect on outflow facility in the normal eyes but caused a 33% decrease in facility in eyes with the GSH-depleted TM. The results indicate that GSH may not participate directly in regulating aqueous humor outflow but is able to protect TM against H202-induced oxidative damage that would otherwise lead to a decrease in outflow facility.
Aqueous humor is known to contain a significant level of H2O2, but the mechanisms by which ocular tissues protect against oxidative damage are not well understood. With the use of C-1-, C-2-, and C-6-labeled glucose, the contribution of glutathione (GSH) metabolism and the hexose monophosphate shunt (HMS) to the detoxification of peroxide in the lens has been evaluated. It was observed that H2O2 in the culture medium disappeared rapidly (0.5 mumol H2O2/lens/hr) upon incubation of a rabbit lens at 37 degrees C. At 0 degrees to 3 degrees C, however, the rate of disappearance of H2O2 was only one fifth of that observed at the higher temperature. In the absence of a lens or after pretreatment of the lens with methyl mercuric hydroxide, the rate of disappearance of peroxide from the medium was reduced to nearly zero. When a nearly constant level of H2O2 (0.05 to 0.07 mM) was maintained in the medium by means of a peristaltic pump, the amount of CO2 liberated by the HMS at 37 degrees C was found to be three times that liberated from lenses cultured in the absence of peroxide. No change was noted in the level of GSH in the H2O2-treated lenses at 37 degrees C. A significant decrease in GSH was observed, however, at 0 degrees to 3 degrees C, suggesting nonenzymatic oxidation of the tripeptide at the lower temperature. The results indicate that GSH metabolism and the HMS pathway contribute significantly to the detoxification of H2O2 in the lens.
UVB radiation from sunlight is known to be a risk factor for human cataract. The purpose in this study was to investigate the ability of a class I UV-blocking soft contact lens to protect against UVB-induced effects on the ocular tissues of the rabbit in vivo.Eyes of rabbits were exposed to UVB light for 30 minutes (270-360 nm, peak at 310 nm, 1.7 mW/cm(2) on the cornea). Eyes were irradiated in the presence of either a UV-blocking senofilcon A contact lens, a minimally UV-blocking lotrafilcon A contact lens, or no contact lens at all. Effects on the cornea and lens were evaluated at various times after exposure.Eyes irradiated with no contact lens protection showed corneal epithelial cell loss plus lens epithelial cell swelling, vacuole formation, and DNA single-strand breaks, as well as lens anterior subcapsular opacification. The senofilcon A lens protected nearly completely against the UVB-induced effects, whereas the lotrafilcon A lens showed no protection.The results indicate that use of a senofilcon A contact lens is beneficial in protecting ocular tissues of the rabbit against the harmful effects of UVB light, including photokeratitis and cataract.
Blood–retinal barrier (BRB) dysfunction represents one of the most significant changes occurring during diabetic retinopathy. We set up a high-reproducible human-based in vitro BRB model using retinal pericytes, retinal astrocytes, and retinal endothelial cells in order to replicate the human in vivo environment with the same numerical ratio and layer order. Our findings showed that high glucose exposure elicited BRB breakdown, enhanced permeability, and reduced the levels of junction proteins such as ZO-1 and VE-cadherin. Furthermore, an increased expression of pro-inflammatory mediators (IL-1β, IL-6) and oxidative stress-related enzymes (iNOS, Nox2) along with an increased production of reactive oxygen species were observed in our triple co-culture paradigm. Finally, we found an activation of immune response-regulating signaling pathways (Nrf2 and HO-1). In conclusion, the present model mimics the closest human in vivo milieu, providing a valuable tool to study the impact of high glucose in the retina and to develop novel molecules with potential effect on diabetic retinopathy.
