Nakagawa Y, Tayama S
Department of Toxicology, Tokyo Metropolitan Research Laboratory of Public Health, Japan.
Arch Toxicol. 2000 Apr;74(2):99-105. doi: 10.1007/s002040050659.
The relation between the metabolism and the cytotoxic effects of bisphenol A (BPA, 2,2-bis(4-hydroxyphenyl)propane) has been studied in freshly isolated rat hepatocytes and isolated hepatic mitochondria. The incubation of hepatocytes with BPA (0.25-1.0 mM) elicited a concentration- and time-dependent cell death, accompanied by losses of intracellular ATP and total adenine nucleotide pools. BPA at a low-toxic level (0.25 mM) in the hepatocyte suspensions was rapidly converted to its major conjugate, BPA-glucuronide, and other minor products without marked loss of cell viability, although at a toxic level (0.5 mM), more than 65% of the compound presented in an unaltered form 2 h after the incubation. Addition of salicylamide (2 mM), non-toxic to hepatocytes during the incubation period, enhanced BPA-induced cytotoxicity and reduced the loss of BPA and the formation of BPA-glucuronide. The addition of BPA to isolated hepatic mitochondria caused a concentration (0-0.5 mM)-dependent increase in the rate of state 4 oxygen consumption in the presence of an FAD-linked substrate (succinate), indicating an uncoupling effect, whereas the rate of state 3 oxygen consumption was inhibited by BPA. Further, the addition of BPA (0.25 mM) reduced state 3 respiration with NAD+-linked substrates (pyruvate plus malate) and/or with the FAD-linked substrate, whereas state 3 respiration with ascorbate plus tetramethyl-p-phenylenediamine (cytochrome oxidase-linked respiration) was not significantly affected by BPA. A comparative study of the toxic effects of BPA and some bisphenols on cell viability (at 1.0 mM) and mitochondrial respiration (at 0.25 mM) revealed that 4,4'-(1,2-diethyl-1,2-ethenediyl)bisphenol (diethylstilbestrol) was more toxic than BPA, followed by 4,4'-methylenediphenol and 4,4'-biphenol. These results indicate that the onset of cytotoxicity caused by BPA may depend on the intracellular energy status and that mitochondria are important targets of the compound. The toxicity caused by the inhibition of ATP synthesis may be related to the concentration of unmetabolised free BPA remaining in the cell suspensions. In addition, the toxic potency of bisphenols to hepatocytes and mitochondria depends on the relative elongation and/or molecular size of the hydrocarbon bridge between the phenolic groups.
已在新鲜分离的大鼠肝细胞和分离的肝线粒体中研究了双酚A（BPA，2,2-双（4-羟基苯基）丙烷）的代谢与其细胞毒性作用之间的关系。用BPA（0.25 - 1.0 mM）孵育肝细胞会引发浓度和时间依赖性的细胞死亡，同时伴随着细胞内ATP和总腺嘌呤核苷酸池的损失。在肝细胞悬液中低毒水平（0.25 mM）的BPA会迅速转化为其主要共轭物BPA - 葡萄糖醛酸苷和其他次要产物，且细胞活力无明显损失，不过在有毒水平（0.5 mM）下，孵育2小时后超过65%的化合物仍以未改变的形式存在。在孵育期间对肝细胞无毒的水杨酰胺（2 mM）的添加增强了BPA诱导的细胞毒性，并减少了BPA的损失以及BPA - 葡萄糖醛酸苷的形成。向分离的肝线粒体中添加BPA会在存在FAD连接底物（琥珀酸）的情况下导致状态4氧气消耗速率出现浓度（0 - 0.5 mM）依赖性增加，表明存在解偶联效应，而状态3氧气消耗速率则受到BPA的抑制。此外，添加BPA（0.25 mM）会降低与NAD⁺连接底物（丙酮酸加苹果酸）和/或与FAD连接底物相关的状态3呼吸作用，而与抗坏血酸加四甲基 - p - 苯二胺相关的状态3呼吸作用（细胞色素氧化酶连接的呼吸作用）未受到BPA的显著影响。一项关于BPA和一些双酚对细胞活力（1.0 mM）和线粒体呼吸作用（0.25 mM）毒性作用的比较研究表明，4,4'-（1,2 - 二乙基 - 1,2 - 乙烯二基）双酚（己烯雌酚）比BPA毒性更大，其次是4,4'-亚甲基双酚和4,4'-联苯酚。这些结果表明，BPA引起的细胞毒性的发生可能取决于细胞内的能量状态，并且线粒体是该化合物的重要靶点。由ATP合成抑制引起的毒性可能与细胞悬液中未代谢的游离BPA的浓度有关。此外，双酚对肝细胞和线粒体的毒性效力取决于酚基团之间烃桥的相对延长和/或分子大小。
