Arteel G E, Thurman R G, Raleigh J A
Department of Pharmacology, University of North Carolina at Chapel Hill, USA.
Eur J Biochem. 1998 May 1;253(3):743-50. doi: 10.1046/j.1432-1327.1998.2530743.x.
2-Nitroimidazoles, such as pimonidazole, are reduced in cells with low oxygen tension and are, therefore, used as hypoxia markers. However, the effect of the pyridine nucleotide redox state on pimonidazole reduction is not known. Therefore, livers from fed or fasted rats were perfused with oxygen-saturated buffer containing pimonidazole (400 microM) in the presence and absence of an inhibitor of the mitochondrial respiratory chain, potassium cyanide; these treatments were used to modulate the mitochondrial and cytosolic pyridine nucleotide redox states. Pimonidazole-induced increases in oxygen uptake over basal values were as follows: fed, 15.1 +/- 2.4; fasted, 4.2 +/- 0.8; fed + KCN, 32.1 +/- 0.9; fasted + KCN, 0.2 +/- 0.2 micromol x g(-1) x h(-1). However, if NADPH was added in excess, microsomal oxygen uptake due to oxidative metabolism of pimonidazole was independent of treatment. These results indicate that pimonidazole-stimulated O2 uptake, due predominantly to N-oxidation and glucuronidation, is dependent on the NADPH redox state. In contrast, reduced pimonidazole adducts, detected immunochemically, accumulated in pericentral regions in liver. Increasing the NADH redox state by inhibiting the mitochondrial respiratory chain with KCN decreased protein-bound pimonidazole adducts. Concomitantly, the average O2 tension of the liver was increased at least 30%. However, KCN had no effect on total pimonidazole adducts detected by ELISA, although both cytosolic (lactate/pyruvate) and mitochondrial (3-hydroxybutyrate/acetoacetate) NADH redox states were elevated by at least a factor of eight. These results indicate that, unlike oxidative metabolism, the pyridine nucleotide redox state does not determine the rate of reductive metabolism of pimonidazole. Instead, the cellular oxygen tension regulates this process. Therefore, even in cases where the supply of reducing equivalents is increased (e.g., ethanol metabolism), accumulation of the reduced bound product of pimonidazole is oxygen dependent in liver.
2-硝基咪唑类化合物，如匹莫硝唑，在低氧张力的细胞中会被还原，因此被用作缺氧标志物。然而，吡啶核苷酸氧化还原状态对匹莫硝唑还原的影响尚不清楚。因此，给喂食或禁食大鼠的肝脏灌注含匹莫硝唑（400微摩尔）的氧饱和缓冲液，同时存在和不存在线粒体呼吸链抑制剂氰化钾；这些处理用于调节线粒体和胞质吡啶核苷酸氧化还原状态。匹莫硝唑诱导的氧摄取相对于基础值的增加如下：喂食组，15.1±2.4；禁食组，4.2±0.8；喂食+氰化钾组，32.1±0.9；禁食+氰化钾组，0.2±0.2微摩尔·克⁻¹·小时⁻¹。然而，如果过量添加烟酰胺腺嘌呤二核苷酸磷酸（NADPH），由于匹莫硝唑的氧化代谢导致的微粒体氧摄取与处理无关。这些结果表明，主要由于N-氧化和葡萄糖醛酸化导致的匹莫硝唑刺激的氧摄取取决于NADPH氧化还原状态。相反，通过免疫化学检测到的还原型匹莫硝唑加合物在肝脏中央周围区域积累。用氰化钾抑制线粒体呼吸链增加烟酰胺腺嘌呤二核苷酸（NADH）氧化还原状态会减少蛋白质结合的匹莫硝唑加合物。同时，肝脏的平均氧张力至少增加了30%。然而，氰化钾对酶联免疫吸附测定（ELISA）检测到的总匹莫硝唑加合物没有影响，尽管胞质（乳酸/丙酮酸）和线粒体（3-羟基丁酸/乙酰乙酸）NADH氧化还原状态都至少升高了8倍。这些结果表明，与氧化代谢不同，吡啶核苷酸氧化还原状态并不决定匹莫硝唑的还原代谢速率。相反，细胞氧张力调节这一过程。因此，即使在还原当量供应增加的情况下（如乙醇代谢），匹莫硝唑还原结合产物在肝脏中的积累也依赖于氧。
