Comparison of primary human hepatocytes and hepatoma cell line HEPG2 with regard to their biotransformation properties
Comparison of primary human hepatocytes and hepatoma cell line HEPG2 with regard to their biotransformation properties
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DOI：
10.1124/dmd.31.8.1035
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发表时间：
2003-08-01
影响因子：
3.9
通讯作者：
Bader, A
中科院分区：
文献类型：
--
作者：
Wilkening, S;Stahl, F;Bader, A
原代肝细胞和肝癌细胞系HepG2的培养常用于人体生物转化研究的体外模型中。在这项研究中，我们对这些模型系统的能力进行了表征和比较，以表明不同类别的诱变剂的存在。对食物诱变剂苯并[a]芘、二甲基亚硝胺和2-amino-1-methyl-6-phenylimidazo[4，5-b]pyridine(PhIP)处理后的遗传毒性敏感性、酶活性和基因表达进行了监测。在夹心培养的原代人肝细胞中，彗星试验可以可靠地检测到DNA损伤。三种诱变剂均可引起原代细胞DNA损伤，但未检测到DMN和PhIP对细胞的遗传毒性作用。我们认为，缺乏特定的酶是他们无法处理这些诱变剂的原因。因此，我们用实时逆转录-聚合酶链式反应对药物代谢酶编码的广泛基因的表达进行了量化。这些基因编码细胞色素P450，此外，还编码一系列重要的II相酶。这些基因在人类肝细胞中的表达水平与之前报道的人类肝脏样本中的表达水平相似。另一方面，在HepG2中的表达水平与人类显著不同。在HepG2细胞中，活性和表达，特别是I相酶的活性和表达极低。在两种细胞类型中，测试物质对特定基因的上调作用是相似的。总之，人肝细胞是人类肝脏生物转化的首选模型，而HepG2细胞可能有助于研究药物代谢酶的调控。
Cultures of primary hepatocytes and hepatoma cell line HepG2 are frequently used in in vitro models for human biotransformation studies. In this study, we characterized and compared the capacity of these model systems to indicate the presence of different classes of promutagens. Genotoxic sensitivity, enzyme activity, and gene expression were monitored in response to treatment with food promutagens benzo[a] pyrene, dimethylnitrosamine (DMN), and 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP). DNA damage could be detected reliably with the comet assay in primary human hepatocytes, which were maintained in sandwich culture. All three promutagens caused DNA damage in primary cells, but in HepG2 no genotoxic effects of DMN and PhIP could be detected. We supposed that the lack of specific enzymes accounts for their inability to process these promutagens. Therefore, we quantified the expression of a broad range of genes coding for drug-metabolizing enzymes with real-time reverse transcription-polymerase chain reaction. The genes code for cytochromes P450 and, in addition, for a series of important phase II enzymes. The expression level of these genes in human hepatocytes was similar to those previously reported for human liver samples. On the other hand, expression levels in HepG2 differed significantly from that in human. Activity and expression, especially of phase I enzymes, were demonstrated to be extremely low in HepG2 cells. Up-regulation of specific genes by test substances was similar in both cell types. In conclusion, human hepatocytes are the preferred model for biotransformation in human liver, whereas HepG2 cells may be useful to study regulation of drug-metabolizing enzymes.