April 2010
Volume 51, Issue 13
Free
ARVO Annual Meeting Abstract  |   April 2010
Inhibition of Retinal Glycolysis by Oxidative Stress. Prevention by Pyruvate
Author Affiliations & Notes
  • K. R. Hegde
    Ophthalmology & Visual Sciences,
    Univ of Maryland Sch of Medicine, Baltimore, Maryland
  • S. Kovtun
    Ophthalmology & Visual Sciences,
    Univ of Maryland Sch of Medicine, Baltimore, Maryland
  • S. D. Varma
    Ophthalmology & Visual Sci & Biochem,
    Univ of Maryland Sch of Medicine, Baltimore, Maryland
  • Footnotes
    Commercial Relationships  K.R. Hegde, None; S. Kovtun, None; S.D. Varma, None.
  • Footnotes
    Support  NIH Grant EY01292
Investigative Ophthalmology & Visual Science April 2010, Vol.51, 1423. doi:
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      K. R. Hegde, S. Kovtun, S. D. Varma; Inhibition of Retinal Glycolysis by Oxidative Stress. Prevention by Pyruvate. Invest. Ophthalmol. Vis. Sci. 2010;51(13):1423.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose: : We have previously shown that pyruvate prevents retinal oxidative stress involved in triggering the pathogenesis of senile macular degeneration, diabetic retinopathy, etc.. Its effect has been previously attributed to its oxyradical scavenging properties. We hypothesize that its protective effect is also attributable to its ability to overcome glycolytic inhibition associated with oxidative stress. The present studies were undertaken to verify this hypothesis.

Methods: : The hypothesis has been verified by measuring glycolysis in retinal explants incubated for 3.5 hrs in medium199 generating ROS with and without pyruvate (10mM) and determining tritiated water (3H2O) generated from 5-3H glucose. ROS was generated by addition of xanthine and xanthine oxidase to the medium. The medium was then analyzed chromatographically for 3H2O. The chromatographic system consisted of an anion exchange (-OH) mini-column piggybacked on a boronate column. The radioactivity in the eluate accounted for 3H2O generated by enolase during glycolysis. Lactate was measured spectrophotometrically by the LDH reaction with NAD as a co-substrate. GSH was measured in the acid extract of the tissue using Ellman’s reaction.

Results: : Incubation with ROS decreased the yield of 3H2O to about 50% (0.77 µmoles/mg protein) of the controls incubated without ROS (1.63µmoles/mg). Pyruvate abolished this decrease. The ROS-induced decrease in lactate as well as GSH were also prevented by pyruvate.

Conclusions: : The results are in conformity with the hypothesis proposed. The mechanism by which pyruvate overcomes ROS induced inhibition of glycolysis is attributable to the prevention of oxidative inactivation of key -SH dependent glycolytic enzymes such as the GAPDH, as well as to its ability to regenerate NAD+ by its conversion to lactate by LDH. Hence the compound protects the tissue against ROS damage by acting as a metabolic agonist as well as an oxyradical scavenger. The findings are considered potentially useful from pharmacological point of view.

Keywords: antioxidants • oxidation/oxidative or free radical damage • metabolism 
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