Hermes-Lima M
Institute of Biochemistry, K. B. Storey Laboratory, Carleton University, Ottawa, Canada.
Free Radic Biol Med. 1995 Sep;19(3):381-90. doi: 10.1016/0891-5849(95)00015-p.
The biosynthetic heme precursor 5-aminolevulinic acid (5-ALA) is a generator of oxygen radicals in vitro and possibly in vivo during pathologic situations of 5-ALA overload, for example, acute intermittent porphyria and saturnism. It has been observed that 5-ALA induces, in isolated rat liver mitochondria, permeabilization of the inner mitochondrial membrane (a phenomenon called permeability transition) as verified by the elimination of the transmembrane electrical potential, Ca2+ release, mitochondrial swelling, and increase in state-4 respiratory rate. The damaging process is primarily attributed to .OH radicals as elucidated by the protection by catalase, superoxide dismutase, and the Fe(II) chelator o-phenanthroline. Ruthenium red, EGTA, and dithiothretol (DTT) have been observed to prevent the action of 5-ALA-generated oxyradicals, suggesting the participation of both Ca2+ and the oxidation of critical thiol membrane proteins in the process of permeability transition. 5-ALA-induced polymerization of thiol membrane proteins has also been demonstrated by SDS-PAGE electrophoresis of the mitochondrial suspensions, a process similar to that observed in mitochondria treated with tert-butyI hydroperoxide. EGTA addition, in contrast with DTT or antioxidants, restores the previously eliminated electrical potential. Furthermore, EGTA prevents the 5-ALA-mediated polimeryzation of thiol proteins. These observations suggest that Ca2+ participates in a later stage of the permeability transition, after the oxidation of the thiol proteins. The effects of 5-ALA-derived oxyradicals in isolated mitochondria could be used as a tool for more general studies of oxidative stress, such as the mitochondrial injury that follows processes of ischemia and reperfusion or xenobiotic poisoning.
生物合成的血红素前体5-氨基乙酰丙酸（5-ALA）在体外是氧自由基的产生剂，在5-ALA过载的病理情况下，如急性间歇性卟啉病和铅中毒，在体内可能也是如此。据观察，5-ALA在离体大鼠肝线粒体中可诱导线粒体内膜通透性转变（一种称为通透性转换的现象），这可通过跨膜电势的消除、Ca2+释放、线粒体肿胀以及状态4呼吸速率的增加来证实。损伤过程主要归因于·OH自由基，这一点已通过过氧化氢酶、超氧化物歧化酶和Fe(II)螯合剂邻菲罗啉的保护作用得以阐明。已观察到钌红、乙二醇双乙醚二胺四乙酸（EGTA）和二硫苏糖醇（DTT）可阻止5-ALA产生的氧自由基的作用，这表明Ca2+和关键巯基膜蛋白的氧化均参与了通透性转换过程。通过线粒体悬浮液的十二烷基硫酸钠-聚丙烯酰胺凝胶电泳（SDS-PAGE）也证实了5-ALA诱导的巯基膜蛋白聚合，这一过程类似于用叔丁基过氧化氢处理的线粒体中观察到的过程。与DTT或抗氧化剂不同，添加EGTA可恢复先前消除的电势。此外，EGTA可阻止5-ALA介导的巯基蛋白聚合。这些观察结果表明，Ca2+在巯基蛋白氧化后参与通透性转换的后期阶段。5-ALA衍生的氧自由基在离体线粒体中的作用可作为一种工具，用于更广泛地研究氧化应激，如缺血再灌注过程或外源化合物中毒后发生的线粒体损伤。
