Dissemination of Peroxidative Stress via Intermembrane Transfer of Lipid Hydroperoxides: Model Studies with Cholesterol Hydroperoxides

Lipid hydroperoxides (LOOHs) can be generated in cells when cholesterol (Ch) and other unsaturated lipids in cell membranes are degraded under conditions of oxidative stress. If LOOHs escape reductive detoxification by glutathione-dependent selenoperoxidases, they may undergo iron-catalyzed one-electron reduction to free radical species, thus triggering peroxidative chain reactions which exacerbate oxidative membrane damage. LOOHs are more polar than parent lipids and much longer-lived than free radical precursors or products. Accordingly, intermembrane transfer of LOOHs (analogous to that of unoxidized precursors) might be possible, and this could jeopardize acceptor membranes. We have investigated this possibility, using photoperoxidized [14C]Ch-labeled erythrocyte ghosts as cholesterol hydroperoxide (ChOOH) donors and unilamellar liposomes [e.g., dimyristoyl-phosphatidylcholine/Ch, 9:1 mol/mol] as acceptors. ChOOH material consisted mainly of 5α-hydroperoxide, a singlet oxygen adduct. Time-dependent transfer of ChOOH versus Ch at 37°C was determined, using high-performance liquid and thin-layer chromatographic methods to analyze liposomal extracts for these species. A typical experiment in which the starting ChOOH/Ch mol ratio in ghosts was ∼0.05 showed that the initial transfer rate of ChOOH was ∼16 times greater than that of parent Ch. Using [14C]Ch as a reporter in liposome acceptors, we found that transfer-acquired ChOOHs, when exposed to a lipophilic iron chelate and ascorbate, could trigger strong peroxidative chain reactions, as detected by accumulation of [14C]Ch oxidation products. These findings support the hypothesis that intermembrane transfer of ChOOHs can contribute to their prooxidant membrane damaging and cytotoxic potential.