Oxidative damage to proteins and lipids of CNS myelin produced by in vitro generated reactive oxygen species

Abstract

Purified myelin isolated from 70-day-old rats was submitted to nonenzymatic peroxidative systems containing 100 microM FeCl3.6H2O, 100 microM ascorbic acid, and 100 microM CuSO4.6H2O 10 mM H2O2 in order to investigate the extent of damage produced by reactive oxygen species (ROS). Iron and copper catalyzing systems were selected because of the known importance of these metals in producing free radical chain reactions in biological membranes (Halliwell and Gutteridge: "Free Radicals in Biology and Medicine," Oxford: Clarendon Press, 1989). Our findings show that: (1) although after 1 hour of peroxidation, an important level of thiobarbituric acid-reactive substances (TBARS) was detected, polyunsaturated fatty acids (20:2; 20:4; 22:4 and 22:6) were markedly affected only after 14 hours of incubation; (2) protein thiol groups were very sensitive to the attack of ROS generated by copper but resistant to iron-generated ROS; (3) aggregation of myelin proteins produced by peroxidation could be prevented by sulfhydryl (SH)-reducing agents, and (4) as a consequence of these modifications, compact myelin suffered disruption of its intraperiodic line. In conclusion, our results demonstrate that this unique membrane of the central nervous system (CNS) is highly vulnerable to oxidative stress and that this susceptibility to oxidative damage could be prevented, at least partially, by the use of SH-protective molecules.