Münzel Thomas, Afanas'ev Igor B, Kleschyov Andrei L, Harrison David G
Universitätsklinik Eppendorf, Hamburg Germany.
Arterioscler Thromb Vasc Biol. 2002 Nov 1;22(11):1761-8. doi: 10.1161/01.atv.0000034022.11764.ec.
During the past decade, it has become apparent that reactive oxygen species play a critical role in the genesis of many vascular diseases. The superoxide anion is among the most important of these, not only because of its rapid reaction with NO but also because it serves as a progenitor for many other reactive oxygen species. Although there are many approaches to detecting and quantifying superoxide in chemical systems, its detection in intact tissues is more difficult. The validity of the most popular and frequently used assay for this purpose, lucigenin-enhanced chemiluminescence, has been recently questioned. It has been suggested that lucigenin itself, especially at high concentrations (>50 micromol/L), may act as a source for superoxide via redox cycling. Lower lucigenin concentrations (5 micromol/L) do not participate in redox cycling to an important extent in intact tissues and, therefore, provide an accurate assessment of the rate of superoxide production in such samples. Other useful assays for superoxide include those using the fluorescent dye dihydroethidine, 2-methyl-6-phenyl-3,7-dihydroimidazo(1,2-alpha)pyrazin-3-one (CLA), and 2-(p-hydroxybenzyl)-6-(p-hydroxyphenyl) 8-benzylimidazo[1,2-alpha]pyrazin-3-one (coelenterazine). The chemiluminescent compound 5-amino-2,3-dihydroxy-1,4-phthalayineidone (luminol) may also be used to detect various reactive oxygen species and may be made specific for various oxidants, such as hydrogen peroxide, superoxide, and peroxynitrite, by altering the experimental conditions. Although each of these methods may be associated with potential artifacts, the use of > or =2 different techniques that yield similar results provides a reliable approach for the study of reactive oxygen species in intact vascular tissues.
在过去十年中，活性氧在许多血管疾病的发生中起着关键作用已变得显而易见。超氧阴离子是其中最重要的之一，这不仅是因为它能与一氧化氮快速反应，还因为它是许多其他活性氧的前体。尽管在化学系统中有许多检测和定量超氧阴离子的方法，但在完整组织中检测它则更为困难。用于此目的的最流行且常用的检测方法，即光泽精增强化学发光法，其有效性最近受到了质疑。有人提出，光泽精本身，尤其是在高浓度（>50微摩尔/升）时，可能通过氧化还原循环充当超氧阴离子的来源。较低的光泽精浓度（5微摩尔/升）在完整组织中在很大程度上不参与氧化还原循环，因此能准确评估此类样品中超氧阴离子的产生速率。其他用于检测超氧阴离子的有用方法包括使用荧光染料二氢乙锭、2-甲基-6-苯基-3,7-二氢咪唑并[1,2-α]吡嗪-3-酮（CLA）以及2-(对羟基苄基)-6-(对羟基苯基)8-苄基咪唑并[1,2-α]吡嗪-3-酮（腔肠素）的方法。化学发光化合物5-氨基-2,3-二羟基-1,4-酞嗪二酮（鲁米诺）也可用于检测各种活性氧，并且通过改变实验条件可使其对各种氧化剂具有特异性，如过氧化氢、超氧阴离子和过氧亚硝酸盐。尽管这些方法中的每一种都可能存在潜在的假象，但使用≥2种产生相似结果的不同技术为研究完整血管组织中的活性氧提供了一种可靠的方法。
