Comparative analysis of 3D culture methods on human HepG2 cells
Comparative analysis of 3D culture methods on human HepG2 cells
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DOI：
10.1007/s00204-016-1677-z
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发表时间：
2017-01-01
影响因子：
6.1
通讯作者：
Hessel, Stefanie
中科院分区：
文献类型：
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作者：
Luckert, Claudia;Schulz, Christina;Hessel, Stefanie
人原代肝细胞是体外肝脏研究的金标准。由于其低可用性和高成本，具有可比形态学和生化特征的替代肝细胞模型已成为焦点。人肝癌细胞系HepG2常被用作毒性研究的肝脏模型。然而，在二维（2D）培养条件下，外源性代谢酶和典型的肝脏标志物如白蛋白的表达非常低。据报道，在三维（3D）Matrigel培养系统中培养21天强烈增加HepG2细胞的代谢能力。在我们目前的研究中，我们进一步比较了HepG2细胞在三种不同的3D系统中的培养：胶原蛋白，Matrigel和Alvetex培养。细胞形态，白蛋白分泌，细胞色素P450单加氧酶的活性，以及外源性代谢和肝脏特异性酶的基因表达后3，7，14，和21天的培养进行了分析。我们的研究结果表明，先前报道的HepG2细胞代谢能力的增加主要不是3D培养的结果，而是培养持续时间的结果。在2D单层中生长21天的HepG2细胞表现出与我们研究中使用的所有3D培养系统相当的生化特性、生物活性和基因表达模式。然而，HepaRG活性没有达到HepaRG细胞的水平。总之，肝癌细胞系HepG2代谢能力的增加不是由于3D培养，而是由于培养时间延长。
Human primary hepatocytes represent a gold standard in in vitro liver research. Due to their low availability and high costs alternative liver cell models with comparable morphological and biochemical characteristics have come into focus. The human hepatocarcinoma cell line HepG2 is often used as a liver model for toxicity studies. However, under two-dimensional (2D) cultivation conditions the expression of xenobiotic-metabolizing enzymes and typical liver markers such as albumin is very low. Cultivation for 21 days in a three-dimensional (3D) Matrigel culture system has been reported to strongly increase the metabolic competence of HepG2 cells. In our present study we further compared HepG2 cell cultivation in three different 3D systems: collagen, Matrigel and Alvetex culture. Cell morphology, albumin secretion, cytochrome P450 monooxygenase enzyme activities, as well as gene expression of xenobiotic-metabolizing and liver-specific enzymes were analyzed after 3, 7, 14, and 21 days of cultivation. Our results show that the previously reported increase of metabolic competence of HepG2 cells is not primarily the result of 3D culture but a consequence of the duration of cultivation. HepG2 cells grown for 21 days in 2D monolayer exhibit comparable biochemical characteristics, CYP activities and gene expression patterns as all 3D culture systems used in our study. However, CYP activities did not reach the level of HepaRG cells. In conclusion, the increase of metabolic competence of the hepatocarcinoma cell line HepG2 is not due to 3D cultivation but rather a result of prolonged cultivation time.