Tyakht Alexander V, Ilina Elena N, Alexeev Dmitry G, Ischenko Dmitry S, Gorbachev Alexey Y, Semashko Tatiana A, Larin Andrei K, Selezneva Oksana V, Kostryukova Elena S, Karalkin Pavel A, Vakhrushev Igor V, Kurbatov Leonid K, Archakov Alexander I, Govorun Vadim M
Research Institute of Physico-Chemical Medicine, Malaya Pirogovskaya 1a, Moscow 119435, Russia.
BMC Genomics. 2014 Dec 15;15(1):1108. doi: 10.1186/1471-2164-15-1108.
Human hepatoma HepG2 cells are used as an in vitro model of the human liver. High-throughput transcriptomic sequencing is an advanced approach for assessing the functional state of a tissue or cell type. However, the influence of experimental factors, such as the sample preparation method and inter-laboratory variation, on the transcriptomic profile has not been evaluated.
The whole-transcriptome sequencing of HepG2 cells was performed using the SOLiD platform and validated using droplet digital PCR. The gene expression profile was compared to the results obtained with the same sequencing method in another laboratory and using another sample preparation method. We also compared the transcriptomic profile HepG2 cells with that of liver tissue. Comparison of the gene expression profiles between the HepG2 cell line and liver tissue revealed the highest variation, followed by HepG2 cells submitted to two different sample preparation protocols. The lowest variation was observed between HepG2 cells prepared by two different laboratories using the same protocol. The enrichment analysis of the genes that were differentially expressed between HepG2 cells and liver tissue mainly revealed the cancer-associated gene signature of HepG2 cells and the activation of the response to chemical stimuli in the liver tissue. The HepG2 transcriptome obtained with the SOLiD platform was highly correlated with the published transcriptome obtained with the Illumina and Helicos platforms, with moderate correspondence to microarrays.
In the present study, we assessed the influence of experimental factors on the HepG2 transcriptome and identified differences in gene expression between the HepG2 cell line and liver cells. These findings will facilitate robust experimental design in the fields of pharmacology and toxicology. Our results were supported by a comparative analysis with previous HepG2 gene expression studies.
人肝癌HepG2细胞用作人类肝脏的体外模型。高通量转录组测序是评估组织或细胞类型功能状态的先进方法。然而，尚未评估诸如样品制备方法和实验室间差异等实验因素对转录组图谱的影响。
使用SOLiD平台对HepG2细胞进行全转录组测序，并使用液滴数字PCR进行验证。将基因表达谱与另一个实验室使用相同测序方法和另一种样品制备方法获得的结果进行比较。我们还将HepG2细胞的转录组图谱与肝组织的转录组图谱进行了比较。HepG2细胞系与肝组织之间基因表达谱的比较显示差异最大，其次是采用两种不同样品制备方案的HepG2细胞。在使用相同方案的两个不同实验室制备的HepG2细胞之间观察到差异最小。对HepG2细胞和肝组织之间差异表达基因的富集分析主要揭示了HepG2细胞的癌症相关基因特征以及肝组织中对化学刺激反应的激活。使用SOLiD平台获得的HepG2转录组与使用Illumina和Helicos平台获得的已发表转录组高度相关，与微阵列有适度的对应关系。
在本研究中，我们评估了实验因素对HepG2转录组的影响，并确定了HepG2细胞系与肝细胞之间基因表达的差异。这些发现将有助于在药理学和毒理学领域进行稳健的实验设计。我们的结果得到了与先前HepG2基因表达研究的比较分析的支持。
