Comparison of coumarin-induced toxicity between sandwich-cultured primary rat hepatocytes and rats in vivo: A toxicogenomics approach
Comparison of coumarin-induced toxicity between sandwich-cultured primary rat hepatocytes and rats in vivo: A toxicogenomics approach
复制标题
DOI：
10.1124/dmd.106.011262
复制
发表时间：
2006-12-01
影响因子：
3.9
通讯作者：
Stierum, Rob H.
中科院分区：
文献类型：
--
作者：
Kienhuis, Anne S.;Wortelboer, Heleen M.;Stierum, Rob H.
夹心培养的原代大鼠肝细胞常用作毒理学和药理学的体外模型。然而，肝脏特异性功能的丧失，特别是细胞色素P450（P450）酶活性的下降，限制了该模型用于预测体内毒性的价值。在这项研究中，我们研究了是否有改善代谢能力的肝脏在体外系统提高香豆素诱导的体内毒性的可预测性，通过使用毒理基因组学方法。因此，原代大鼠肝细胞在含有低浓度P450诱导剂、苯巴比妥、地塞米松和β-萘醌的混合物的培养基中以夹心结构培养。所使用的毒理基因组学方法能够在体外和体内条件下在分子水平上比较相似的机制终点，即化合物诱导的多个基因和信号传导途径的变化。将在改良培养基中培养的肝细胞和在标准培养基（不含诱导剂）中培养的肝细胞中观察到的毒性诱导的细胞毒性效应和基因表达谱与大鼠体内研究的结果进行比较。香豆素被用作模型化合物，因为其毒性取决于P450酶的生物活化。与标准培养基中培养的肝细胞相比，改良培养基中培养的肝细胞中香豆素向活性代谢物的代谢、香豆素诱导的细胞毒性和基因表达调节更为明显。此外，在改良培养基中培养的肝细胞和体内培养的肝细胞中，香豆素对更多基因和生物学途径的影响相似。总之，这些实验表明，对于香豆素诱导的毒性，与在标准培养基中培养的肝细胞相比，在改良培养基中维持的体外培养的肝细胞更好地代表了体内情况。
Sandwich-cultured primary rat hepatocytes are often used as an in vitro model in toxicology and pharmacology. However, loss of liver-specific functions, in particular, the decline of cytochrome P450 (P450) enzyme activity, limits the value of this model for prediction of in vivo toxicity. In this study, we investigated whether a hepatic in vitro system with improved metabolic competence enhances the predictability for coumarin-induced in vivo toxicity by using a toxicogenomics approach. Therefore, primary rat hepatocytes were cultured in sandwich configuration in medium containing a mixture of low concentrations of P450 inducers, phenobarbital, dexamethasone, and beta-naphthoflavone. The toxicogenomics approach used enabled comparison of similar mechanistic end-points at the molecular level between in vitro and in vivo conditions, namely, compound-induced changes in multiple genes and signaling pathways. Toxicant-induced cytotoxic effects and gene expression profiles observed in hepatocytes cultured in modified medium and hepatocytes cultured in standard medium (without inducers) were compared with results from a rat in vivo study. Coumarin was used as a model compound because its toxicity depends on bioactivation by P450 enzymes. Metabolism of coumarin toward active metabolites, coumarin-induced cytotoxicity, and gene expression modulation were more pronounced in hepatocytes cultured in modified medium compared with hepatocytes cultured in standard medium. In addition, more genes and biological pathways were similarly affected by coumarin in hepatocytes cultured in modified medium and in vivo. In conclusion, these experiments showed that for coumarin-induced toxicity, sandwich-cultured hepatocytes maintained in modified medium better represent the situation in vivo compared with hepatocytes cultured in standard medium.