Porubek D J, Rundgren M, Harvison P J, Nelson S D, Moldéus P
Mol Pharmacol. 1987 Jun;31(6):647-53.
The toxicity of acetaminophen (4'-hydroxyacetanilide), 3,5-dimethylacetaminophen (3'-5'-dimethyl-4'-hydroxyacetanilide), and 2,6-dimethylacetaminophen (2',6'-dimethyl-4'-hydroxyacetanilide) was investigated in hepatocytes isolated from phenobarbital-pretreated rats. At a concentration of 5 mM, acetaminophen was found to be the most cytotoxic of the three analogues. Inhibition of cellular glutathione reductase by pretreatment of hepatocytes with BCNU enhanced the toxicity of 3,5-dimethylacetaminophen without affecting the toxicity of either acetaminophen or 2,6-dimethylacetaminophen. In contrast, pretreatment with diethylmaleate preferentially enhanced the toxicity caused by 2,6-dimethylacetaminophen and, to a lesser extent, acetaminophen, without measurably affecting the toxicity of 3,5-dimethylacetaminophen. All three hydroxyacetanilides depleted cellular glutathione concentrations, but only the 3,5-dimethyl analogue caused measurable formation of glutathione disulfide. However, the cytotoxicity of all analogues could be decreased by the administration of the thiol agent, dithiothreitol. Moreover, all three analogues had antioxidant properties, and their ability to decrease cellular malondialdehyde formation correlated with their half-wave (E1/2) oxidation potentials. The administration of the ferric ion chelator, desferrioxamine, which completely inhibited lipid peroxidation as measured by malondialdehyde formation, had no significant effects on cytotoxicity caused by acetaminophen or 3,5-dimethylacetaminophen, but partially protected against cytotoxicity caused by 2,6-dimethylacetaminophen, the poorest antioxidant of the three analogues. Covalent protein binding of all three analogues was measured. Whereas both acetaminophen and 2,6-dimethylacetaminophen bound to hepatocyte proteins under conditions where they were cytotoxic, 3,5-dimethylacetaminophen did not. Dithiothreitol was found to decrease the binding of radiolabel from both acetaminophen and its 2,6-dimethyl analogue, whereas desferrioxamine had no effect. These data indicate that the three analogues cause their cytotoxic effects by different mechanisms, although toxicity in all cases is probably mediated through their oxidation products, the quinone imines, which have as a common feature their ability to deplete cellular thiols.
对从经苯巴比妥预处理的大鼠分离出的肝细胞，研究了对乙酰氨基酚（4'-羟基乙酰苯胺）、3,5-二甲基对乙酰氨基酚（3'-5'-二甲基-4'-羟基乙酰苯胺）和2,6-二甲基对乙酰氨基酚（2',6'-二甲基-4'-羟基乙酰苯胺）的毒性。在浓度为5 mM时，发现对乙酰氨基酚是这三种类似物中细胞毒性最大的。用卡莫司汀预处理肝细胞抑制细胞谷胱甘肽还原酶，增强了3,5-二甲基对乙酰氨基酚的毒性，而不影响对乙酰氨基酚或2,6-二甲基对乙酰氨基酚的毒性。相反，用马来酸二乙酯预处理优先增强了2,6-二甲基对乙酰氨基酚以及在较小程度上对乙酰氨基酚所引起的毒性，而对3,5-二甲基对乙酰氨基酚的毒性没有明显影响。所有三种羟基乙酰苯胺都降低了细胞内谷胱甘肽浓度，但只有3,5-二甲基类似物导致了可测量的谷胱甘肽二硫化物的形成。然而，所有类似物的细胞毒性都可以通过给予硫醇试剂二硫苏糖醇而降低。此外，所有三种类似物都具有抗氧化特性，它们降低细胞丙二醛形成的能力与其半波（E1/2）氧化电位相关。给予铁离子螯合剂去铁胺，它能完全抑制以丙二醛形成来衡量的脂质过氧化，对由对乙酰氨基酚或3,5-二甲基对乙酰氨基酚引起的细胞毒性没有显著影响，但能部分保护细胞免受2,6-二甲基对乙酰氨基酚（这三种类似物中抗氧化能力最差的）引起的细胞毒性。测定了所有三种类似物的共价蛋白结合情况。在对乙酰氨基酚和2,6-二甲基对乙酰氨基酚具有细胞毒性的条件下，它们都能与肝细胞蛋白结合，而3,5-二甲基对乙酰氨基酚则不能。发现二硫苏糖醇能降低对乙酰氨基酚及其2,6-二甲基类似物的放射性标记结合，而去铁胺则没有作用。这些数据表明，这三种类似物通过不同机制引起细胞毒性，尽管在所有情况下毒性可能都是通过它们的氧化产物醌亚胺介导的，醌亚胺的一个共同特征是它们能够消耗细胞内的硫醇。
