The primary objective of the proposal is to study biochemical and pharmacological strategies for preventing cataractogenesis using diabetic and oxidative stress animal models and lens culture. We anticipate that certain physiological alpha keto acids could play such a preventive role, especially pyruvate. This is based on our observation that it inhibits polyol synthesis, glycation, AGE formation, and oxidative stress in lens culture and most importantly, its effectiveness against the actual cataract formation in galactosemic animals. We will now test the hypotheses (I) that exogenously administered pyruvate will inhibit cataractogenesis in diabetic rats with high lens aldose reductase (AR) activity, (2) that it will be effective also in diabetic mice with low lens AR, but being subject to ROS and glycation stress, causing structural and functional protein and lipid changes, (3) that the chaperone activity of the alpha-crystallin is decreased by the high levels of fructose, as in the diabetic lens, and (4) that pyruvate will also protect against the combined stress of diabetes and oxidation. The pyruvate effects are attributable to the reactivity of its keto group, situated adjacent to the carboxyl group. Therefore, we hypothesize (5) that other keto acids with greater cell permeability and lower metabolism should be physiologically more effective. Hypotheses #1 to #4 will be verified by the delay in cataract formation in diabetic animals given pyruvate in the diet and drinking water with reference to the controls with comparable hyperglycemia. Basal controls with and without pyruvate will also be run. GSH-peroxidase knockout mice will be used for specific aim #3. The levels of sorbitol, fructose, glycated proteins, AGE, ATP, GSH, malonaldehyde (MDA) and the chaperone activity of alpha-crystallin will be determined in all the groups. Hypothesis #5 will be verified by the relative activity of the keto acids in scavenging ROS, and their effectiveness in preventing lens damage in culture, as indexed by the cation pump activity, and the above biochemical parameters.
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