Uehara T, Kiyosawa N, Shimizu T, Omura K, Hirode M, Imazawa T, Mizukawa Y, Ono A, Miyagishima T, Nagao T, Urushidani T
Toxicogenomics Project, National Institute of Biomedical Innovation, Ibaraki, Osaka, Japan.
Hum Exp Toxicol. 2008 Jan;27(1):23-35. doi: 10.1177/0960327107087910.
One expected result from toxicogenomics technology is to overcome the barrier because of species-specific differences in prediction of clinical toxicity using animals. The present study serves as a model case to test if the well-known species-specific difference in the toxicity of coumarin could be elucidated using comprehensive gene expression data from rat in-vivo, rat in-vitro, and human in-vitro systems. Coumarin 150 mg/kg produced obvious pathological changes in the liver of rats after repeated administration for 7 days or more. Moreover, 24 h after a single dose, we observed minor and transient morphological changes, suggesting that some early events leading to hepatic injury occur soon after coumarin is administered to rats. Comprehensive gene expression changes were analyzed using an Affymetrix GeneChip approach, and differentially expressed probe sets were statistically extracted. The changes in expression of the selected probe sets were further examined in primary cultured rat hepatocytes exposed to coumarin, and differentially expressed probe sets common to the in-vivo and in-vitro datasets were selected for further study. These contained many genes related to glutathione metabolism and the oxidative stress response. To incorporate human data, human hepatocyte cultured cells were exposed to coumarin and changes in expression of the bridging gene set were examined. In total, we identified 14 up-regulated and 11 down-regulated probe sets representing rat-human bridging genes. The overall responsiveness of these genes to coumarin was much higher in rats than humans, consistent with the reported species difference in coumarin toxicity. Next, we examined changes in expression of the rat-human bridging genes in cultured rat and human hepatocytes treated with another hepatotoxicant, diclofenac sodium, for which hepatotoxicity does not differ between the species. Both rat and human hepatocytes responded to the marker genes to the same extent when the same concentrations of diclofenac sodium were exposed. We conclude that toxicogenomics-based approaches show promise for overcoming species-specific differences that create a bottleneck in analysis of the toxicity of potential therapeutic treatments.
毒理基因组学技术的一个预期成果是克服因使用动物预测临床毒性时存在物种特异性差异而产生的障碍。本研究作为一个模型案例，以检验是否可以利用来自大鼠体内、大鼠体外和人类体外系统的全面基因表达数据来阐明香豆素毒性中众所周知的物种特异性差异。香豆素150mg/kg连续给药7天或更长时间后，大鼠肝脏出现明显的病理变化。此外，单次给药24小时后，我们观察到轻微且短暂的形态学变化，这表明在香豆素给大鼠给药后不久就会发生一些导致肝损伤的早期事件。使用Affymetrix基因芯片方法分析全面的基因表达变化，并统计提取差异表达的探针集。在暴露于香豆素的原代培养大鼠肝细胞中进一步检测所选探针集的表达变化，并选择体内和体外数据集中共有的差异表达探针集进行进一步研究。这些探针集包含许多与谷胱甘肽代谢和氧化应激反应相关的基因。为纳入人类数据，将人类肝细胞培养细胞暴露于香豆素，并检测桥接基因集的表达变化。我们总共鉴定出14个上调和11个下调的代表大鼠 - 人类桥接基因的探针集。这些基因对香豆素的总体反应性在大鼠中比在人类中高得多，这与报道的香豆素毒性的物种差异一致。接下来，我们检测了用另一种肝毒性药物双氯芬酸钠处理的培养大鼠和人类肝细胞中大鼠 - 人类桥接基因的表达变化，双氯芬酸钠的肝毒性在不同物种之间没有差异。当暴露于相同浓度的双氯芬酸钠时，大鼠和人类肝细胞对标记基因的反应程度相同。我们得出结论，基于毒理基因组学的方法有望克服物种特异性差异，而这种差异在潜在治疗药物毒性分析中造成了瓶颈。
