Vila A, Korytowski W, Girotti A W
Department of Biochemistry, Medical College of Wisconsin, Milwaukee, Wisconsin 53226, USA.
Arch Biochem Biophys. 2000 Aug 1;380(1):208-18. doi: 10.1006/abbi.2000.1928.
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 [(14)C]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 5alpha-hydroperoxide, a singlet oxygen adduct. Time-dependent transfer of ChOOH versus Ch at 37 degrees 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 approximately 0.05 showed that the initial transfer rate of ChOOH was approximately 16 times greater than that of parent Ch. Using [(14)C]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 [(14)C]Ch oxidation products. These findings support the hypothesis that intermembrane transfer of ChOOHs can contribute to their prooxidant membrane damaging and cytotoxic potential.
当细胞膜中的胆固醇（Ch）和其他不饱和脂质在氧化应激条件下被降解时，细胞中会生成脂质氢过氧化物（LOOHs）。如果LOOHs未能通过谷胱甘肽依赖性硒过氧化物酶进行还原性解毒，它们可能会经历铁催化的单电子还原生成自由基，从而引发过氧化链式反应，加剧氧化膜损伤。LOOHs比母体脂质极性更强，寿命也比自由基前体或产物长得多。因此，LOOHs（类似于未氧化的前体）的膜间转移可能是可行的，这可能会危及受体膜。我们利用光过氧化的[(14)C]Ch标记红细胞膜作为胆固醇氢过氧化物（ChOOH）供体，单层脂质体[如二肉豆蔻酰磷脂酰胆碱/Ch，9:1摩尔/摩尔]作为受体，研究了这种可能性。ChOOH物质主要由5α-氢过氧化物（一种单线态氧加合物）组成。在37℃下，使用高效液相色谱和薄层色谱方法分析脂质体提取物中的这些物质，测定了ChOOH与Ch随时间的转移情况。一个典型实验中，膜中起始ChOOH/Ch摩尔比约为0.05，结果表明ChOOH的初始转移速率约为母体Ch的16倍。在脂质体受体中使用[(14)C]Ch作为报告物，我们发现转移获得的ChOOHs在暴露于亲脂性铁螯合剂和抗坏血酸时，可引发强烈的过氧化链式反应，这可通过[(14)C]Ch氧化产物的积累检测到。这些发现支持了以下假设：ChOOHs的膜间转移可导致其促氧化膜损伤和细胞毒性潜力。
