Abstract

Introduction: Aberrations in retinal physiology, metabolism, and morphology, are known to occur due to oxidative stress, a key factor in the pathogenesis of several retinal diseases such as age-related macular degeneration (AMD) and diabetic retinopathy (DR). Functionally, such alterations translate to progressive vision impairment and, if severe enough, may even lead to blindness. We have previously shown that biochemical and metabolic dysfunction, specifically glycolysis, in the retina due to oxyradical exposure can be prevented by pyruvate, an effective scavenger of reactive oxygen species (ROS). This study was undertaken to investigate the mechanism by which pyruvate prevented inhibition of glycolysis.

Method: Bovine neural retinas were incubated in medium 199 +/-ROS and +/- sodium pyruvate (SP) for 6 hours. Xanthine and xanthine oxidase served as the source of ROS. The activities of crucial glycolytic enzymes glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and pyruvate kinase (PK) were determined at 2-hour and 6-hour time points. ATP levels were also determined.

Result: Exposure to ROS resulted in a ~30% decrease in GAPDH activity after 2 hours in both ROS and ROS + pyruvate groups, compared to the controls. This decrease was almost completely reversed after 6 hours of incubation in the ROS group, while the lowered activity persisted in the ROS + pyruvate group. After 2-hours, a 55% decrease occurred in PK activity in the ROS group compared to controls, while only ~10% decrease was observed in ROS+ pyruvate group. At 6-hour time point a sustained 50% decrease in PK activity was seen in the ROS-exposed retinas; in the ROS + pyruvate group it was maintained at ~100% of the control levels. ATP levels were also found to follow a trend similar to PK activity.

Conclusion: Results suggest that sodium pyruvate prevents inactivation of pyruvate kinase induced by ROS; this has been demonstrated for the first time, especially in the retina. The substantially higher level of ATP in the pyruvate group indicates that pyruvate supplementation increases the flux of intermediates through glycolysis by preventing oxidative inactivation of PK. Increasing the activity of Krebs cycle could be another reason for this observation. Results indicate that pyruvate is a promising novel neuroprotective compound, with implications on its pharmacological applications in oxidative stress-involving retinal diseases such as AMD and DR.