Antioxidants Prevention of Diabetic Damage in the Organ Culture Bovine Lenses

Abstract

Oxidative stress represents a mechanism which could lead to diabetic cataract. We exposed bovine lenses in culture conditions for two weeks to high glucose concentration (450 mg%) and investigated the damage to the lens and possible protection by special antioxidants -N-acetyl-L-cysteine (NAC) and the zinc complex of desferrioxamine (DFO), a selective chelator for iron. We monitored the optical quality of the lenses and the oxidation of the epithelium with dichlorofluorescein (DCF) assay, as well as the changes in lens proteins profile by 2D gel electrophoresis. Under high glucose changes in lens focal length, increased oxidation, and changes in lens crystalline were observed. NAC and Zn-DFO nearly completely protected the lenses; DFO showed only partial protection. The results demonstrated that antioxidants should be considered as treatment modality protecting the lens from high glucose damage. It is proposed that a combination of NAC and Zn/DFO could prove highly efficient.the presence of high glucose concentrations (450 mg %), mimicking the diabetic state, and examined the protection bestowed by the DFO complexes and NAC, on the injurious processes on the lenses. We used a unique system of intact bovine lenses, maintained for a longterm, under culture conditions. This system allows for direct exposure to a pre-set glucose concentration, and monitoring the effects on lens transparency with a highly sensitive optical method [20]. The system can detect early optical damage to lenses, which cannot be detected by other methods often used. At the completion of the culture period lens epithelium and lens proteins were analyzed.It was anticipated that the extent of protection exerted on the lens, in this system, will be indicative of the involvement of iron as a catalyst and of free radicals as causative agents, in the damage to the diabetic lens. Materials and Methods Lens organ culture systemLenses were excised in a delicate operation from eyes obtained from 1-year-old male calves under sterile conditions, 2-4 hours after enucleating. From each animal one eye is used for experimental treatment and the other eye serves as control. Each lens is placed in a specially designed culture container, which we have developed [21]. The culture medium consists of M199 with Earl's balanced salt solution, supplemented with 5.96g/L HEPES, 3% dialyzed fetal calf serum and antibiotics (penicillin 100 U/ml and streptomycin 0.1 mg/ ml). Our intact lens culture system mimics the lens conditions inside the eye and makes it possible to keep lenses for long-term studies for