Oxidative modification of rat eye lens proteins by peroxyl radicals in vitro: Protection by the chain-breaking antioxidants stobadine and Trolox

Abstract

In an attempt to model the processes of free radical-mediated cataractogenesis, we investigated the oxidative modification of rat eye lens proteins by peroxyl radicals generated by thermal decomposition of 2,2'-azobis(2-amidinopropane)hydrochloride (AAPH) under aerobic conditions. When incubated with AAPH, the soluble eye lens proteins precipitated in a time-dependent manner. The insolubilisation was accompanied by the accumulation of protein free carbonyls and the diminution of sulfhydryls, yet the processes were shifted in time. The SDS-PAGE analysis of the AAPH-treated proteins revealed the presence of high molecular weight cross-links and, to a lesser extent, fragments. The aggregation and cross-linking of proteins along with the generation of free carbonyls was significantly inhibited by the chain-breaking antioxidants stobadine and Trolox. On the other hand, the AAPH-initiated sulfhydryl consumption was much less sensitive to the antioxidants studied. The results point to a complex mechanism of peroxyl-radical-mediated modification of eye lens proteins with implications for cataract development and they indicate a potentially protective role of antioxidants.

Stobadine – An Indole Type Alternative to the Phenolic Antioxidant Reference Trolox

“…Trolox, in comparison with stobadine, was found to be a more potent inhibitor of 2,2'-azobis-2-amidinopropane (AAPH)-induced precipitation of the soluble eye lens proteins (Stefek et al, 2005). In contrary, production of free carbonyls due to protein oxidation was more efficiently inhibited by stobadine.…”

Vitamin E Analogue Protects Red Blood Cells against Storage-Induced Oxidative Damage

“…Studies by Regoli et al [38] on the capacities of antioxidants to scavenge different radicals demonstrated that mannitol had a much higher scavenging ability towards hydroxyl radicals, generated by the iron/ascorbate-driven Fenton reaction, than towards peroxyl radicals or peroxynitrite. Trolox has been reported to efficiently scavenge a wide variety of free radicals, including the hydroxyl radical and bioradical products arising from · OH radical interactions [39,40]. Similar to α-tocopherol, Trolox is primarily a chain-breaking antioxidant.…”

‘In vitro’ Protective Effect of a Hydrophilic Vitamin E Analogue on the Decrease in Levels of Elongation Factor 2 in Conditions of Oxidative Stress

“…To test whether these alterations in the eEF-2 molecules could be prevented by vitamin E, we studied the effect of Trolox. The antioxidant activity of Trolox has previously been described [43,44] . The idea was also to establish the possible involvement of lipoperoxyl radicals in the mechanism of damage.…”