Stewart R K, Serabjit-Singh C J, Massey T E
Department of Pharmacology and Toxicology, Queen's University, Kingston, Ontario, Canada.
Toxicol Appl Pharmacol. 1996 Oct;140(2):499-507. doi: 10.1006/taap.1996.0246.
Aflatoxin B1 (AFB1) requires bioactivation to AFB1-8,9-epoxide for carcinogenicity, and glutathione S-transferase (GST)-catalyzed conjugation of activated AFB1 with glutathione (GSH) is a critical determinant of susceptibility to the mycotoxin. Incubations containing [3H]AFB1, rabbit liver microsomes, an NADPH-generating system, 1 mM GSH, and GST-containing lung or liver cytosol were performed to assess the abilities of lung and liver GSTs to conjugate AFB1-8,9-epoxide. [3H]AFB1-GSH was isolated by isocratic reverse-phase high-performance liquid chromatography (HPLC) and quantitated by liquid scintillation spectroscopy. Maximal [3H]AFB1-GSH formation rates were significantly lower for lung than for liver (0.3 +/- 0.1 and 1.7 +/- 0.4 nmol/mg/hr, respectively). Immunoprecipitation of rabbit pulmonary cytosolic GSTs with anti-alpha or anti-mu GST antisera decreased [3H]AFB1-GSH production by approximately 45 and 51%, respectively, indicating that alpha-class and mu-class GSTs are of similar importance in catalyzing this reaction in the lung. Because mu-class GSTs comprise only a small proportion of total lung GST content, these enzymes have high specific activity toward AFB1-8,9-epoxide. In contrast, the pi-class GST appeared to play a negligible role. Using a rat liver microsomal system to generate both AFB1 exo- and endoepoxide isomers, and analysis based on chiral HPLC, we found that rabbit liver cytosolic GSTs catalyzed formation of both AFB1 exo- and endo-epoxide-GSH conjugates, whereas pulmonary cytosolic GSTs catalyzed formation of only the exo stereoisomer at detectable levels. Despite a preference for conjugating the more mutagenic AFB1 exo-epoxide isomer, the relatively low capacity for GST-catalyzed detoxification of bioactivated AFB1 in lung may be an important factor in the susceptibility of the lung to AFB1 toxicity.
黄曲霉毒素B1（AFB1）需经生物活化生成AFB1-8,9-环氧化物才具有致癌性，而谷胱甘肽S-转移酶（GST）催化活化的AFB1与谷胱甘肽（GSH）结合是决定对该霉菌毒素易感性的关键因素。进行了含有[3H]AFB1、兔肝微粒体、一个NADPH生成系统、1 mM GSH以及含GST的肺或肝细胞溶胶的孵育实验，以评估肺和肝GST将AFB1-8,9-环氧化物结合的能力。通过等度反相高效液相色谱（HPLC）分离[3H]AFB1-GSH，并通过液体闪烁光谱法进行定量。肺中[3H]AFB1-GSH的最大生成速率显著低于肝脏（分别为0.3±0.1和1.7±0.4 nmol/mg/小时）。用抗α或抗μ GST抗血清对兔肺细胞溶胶GST进行免疫沉淀，分别使[3H]AFB1-GSH的产生减少约45%和51%，表明α类和μ类GST在肺中催化该反应具有相似的重要性。由于μ类GST仅占肺GST总含量的一小部分，这些酶对AFB1-8,9-环氧化物具有高比活性。相比之下，π类GST似乎起的作用可忽略不计。使用大鼠肝微粒体系统生成AFB1外消旋和内消旋环氧化物异构体，并基于手性HPLC进行分析，我们发现兔肝细胞溶胶GST催化生成AFB1外消旋和内消旋环氧化物-GSH共轭物，而肺细胞溶胶GST仅催化生成可检测水平的外消旋立体异构体。尽管更倾向于结合致突变性更强的AFB1外消旋环氧化物异构体，但肺中GST催化生物活化的AFB1解毒的能力相对较低，这可能是肺对AFB1毒性易感性的一个重要因素。
