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S.D. Varma, K. Hegde, S. Kovtun; Peroxynitrite Induced Decarboxylation of Pyruvate – Implications in the Prevention of –SH Oxidation and Lens Damage by Nitric Oxide Derivatives . Invest. Ophthalmol. Vis. Sci. 2006;47(13):3519.
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© ARVO (1962-2015); The Authors (2016-present)
Oxygen centered free radicals have been proposed to make a significant contribution to cataractogenesis, including diabetic cataracts. A number of studies show that nitric oxide, another free radical species, also increases in diabetic aqueous, with bearing on the pathogenesis of cataracts. The active agent in this case could be NO itself or many of its subsequent derivatives in various oxidation states including peroxynitrite. The present investigations were designed to study the possibility of decarboxylation of pyruvate (PY) by such derivatives, specially by peroxynitrite, one of the most potent NO derivative.
Decarboxylation of PY was studied by incubating it with SIN–1 and determining CO2 generation. Briefly, Sodium PY in amounts ranging from 0 to 5 µmoles mixed with 1–C–14 labeled PY acting as a tracer was incubated with 1 µmole of SIN–1 for a period of 40 minutes at 37 º in bicarbonate buffer, pH 7.8. The CO2 generated was then determined by its distillation after acidification of the reaction mixture and trapping it in Hyamine Hydroxide. A study of the possible protection of the tissue thiols against nitrosative oxidation was done by incubation of the supernatant of 0.1% rat lens homogenate containing 0.035 mM –SH with 0.4 mM SIN–1 for 15 minutes at 37º, without or with PY (10mM) and then determining the final thiol levels. Subsequently the levels of acid soluble thiols were also determined. Basic experiments were done with pure GSH.
The decarboxylation of PY by SIN–1 was found to be stoichiometric till 0.6 µmoles, the reaction becoming asymptotic subsequently. Additionally, the concentration of –SH in the lens centrifugates used as above also decreased from the initial level of 0.035 mM to 0.014 mM in 15 minutes in presence of SIN–1. This decrease was also substantially abolished by PY. Similar protective effects of pyruvate were obtained in experiments with pure GSH as well as with acid soluble lens thiols.
The results demonstrate that pyruvate scavenges peroxynitrite by undergoing stochiometric decarboxylation. Consequently, it also inhibits tissue thiol depletion by nitrosation and oxidation, caused by generation of nitric oxide and its conversion to peroxynitrite. The findings are consistent with the hypothesis that pyruvate is helpful in offsetting tissue toxicity caused by availability of excessive levels of NO and other nitric oxide based toxic derivatives, in addition to its properties of protecting the lens against the toxicity of oxygen centered free radicals and high levels of sugars.
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