Stimulation of Glycolysis in the Lens by Pyruvate. Implications in Protection against Oxidative Stress

Abstract

Purpose: We have previously demonstrated that pyruvate protects the lens and other ocular tissues from oxidative stress. This effect is attributable to it action as A potent scavenger of reactive species of oxygen (ROS). The reactions are associated with its decarboxylation, generating acetate, CO2, and H2O, all being relatively innocuous compared to the products of the reaction of ROS with many other antioxidants. The aim of this study was to ascertain that, in addition to the ROS-scavenging action, its protective effect could also be exerted and augmented by its ability to stimulate glycolysis, via an acceleration of NAD+/NADH cycling, maintaining A continued supply of NAD+. Methods: The hypothesis was investigated by incubating mice lenses in Tyrode buffer containing 1 mM glucose and mixed with 5-3H-glucose as A tracer, without/with 2mM sodium pyruvate. The extent of glycolysis achieved, indicated by the amount of 3H2O and 3H-lactate generated, was quantified by liquid scintillation counting after separating them by piggy backed ion exchange (formate column)and affinity (boronate column) chromatography. Results: As expected, the generation of both these compounds was significantly elevated by pyruvate. The amount of 3H2O generated in presence of pyruvate was ~87 nmoles/lens-substantially higher as compared to that in its absence, wherein it was only ~43 nmoles/lens. 3H-lactate generated in the presence of pyruvate was also significantly higher, ~6.5 nmoles/lens vs. ~4.2 nmoles/lens in its absence. ATP levels were also higher with pyruvate. Conclusions: The results demonstrating an increase in formation of 3H2O by lenses incubated with 5-3H glucose in presence of pyruvate, with the concomitant increase in the contents of 3H-lactate, strongly suggest that pyruvate enhances glycolysis. This is attributable to the reduction of pyruvate to lactate by lactate dehydrogenase, coupled with generation of NAD+ from NADH; the NAD+ in turn being re-used to drive the glyceraldehyde-3-phosphate dehydrogenase reaction further down in glycolysis, with the consequence of increased ATP formation. The results, hence, strongly indicate that the protective effect of pyruvate against oxidative stress to the lens and consequent cataract formation is multimodal, involving deactivation of the ROS as well as its action as A metabolic agonist.