Human liver cell line HepaRG is a well-known source of human hepatocyte-like cells which, however, displays limited biotransformation and a tendency to transform after 20 passages. The new HepaRG-CAR cell line overexpressing constitutive androstane receptor (CAR, NR1I3), a regulator of detoxification and energy metabolism outperforms the parental HepaRG cell line in various liver functions. To further characterize this cell line and assess its stability we compared HepaRG-CAR with HepaRG cells at different passages for their expression profile, ammonia and lactate metabolism, bile acid and reactive oxygen species (ROS) production. Transcriptomic profiling of HepaRG-CAR vs. HepaRG early-passage revealed downregulation of hypoxia, glycolysis and proliferation and upregulation of oxidative phosphorylation genesets. In addition CAR overexpression downregulated the mTORC1 signaling pathway, which, as mediator of proliferation and metabolic reprogramming, may play an important role in the establishment of the HepaRG-CAR phenotype. The ammonia and lactate metabolism and bile acid production of HepaRG-CAR cells was stable for 10 additional passages compared to HepaRG cells. Interestingly, bile acid production was 4.5-fold higher in HepaRG-CAR vs. HepaRG cells, whereas lactate and ROS production were 2.7- and 2.0-fold lower, respectively. Principal component analysis showed clustering of HepaRG-CAR (early- and late-passage) and HepaRG early-passage and not with HepaRG late-passage indicating that passaging exerted larger effect on the transcriptional profile of HepaRG than HepaRG-CAR cells. In conclusion, overexpression of CAR in HepaRG cells improves their bile acid production, mitochondrial energy metabolism, and stability, with the latter possibly due to reduced ROS production, resulting in an optimized source of human hepatocytes.

中文翻译：

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全基因组表达谱揭示了人肝细胞系 HepaRG-CAR 的稳定性和线粒体能量代谢增加。


人肝细胞系HepaRG是众所周知的人肝细胞样细胞来源，然而，其生物转化能力有限，并且在传代20次后有转化的趋势。新的HepaRG-CAR细胞系过表达组成型雄甾烷受体（CAR，NR1I3），一种解毒和能量代谢的调节剂，在各种肝功能方面优于亲代HepaRG细胞系。为了进一步表征该细胞系并评估其稳定性，我们比较了 HepaRG-CAR 与不同传代的 HepaRG 细胞的表达谱、氨和乳酸代谢、胆汁酸和活性氧 (ROS) 的产生。 HepaRG-CAR 与 HepaRG 早期传代的转录组分析揭示了缺氧、糖酵解和增殖的下调以及氧化磷酸化基因组的上调。此外，CAR 过表达下调了 mTORC1 信号通路，该通路作为增殖和代谢重编程的介质，可能在 HepaRG-CAR 表型的建立中发挥重要作用。与 HepaRG 细胞相比，HepaRG-CAR 细胞的氨和乳酸代谢以及胆汁酸生成在额外 10 次传代中保持稳定。有趣的是，HepaRG-CAR 细胞的胆汁酸产量比 HepaRG 细胞高 4.5 倍，而乳酸和 ROS 产量分别低 2.7 倍和 2.0 倍。主成分分析显示HepaRG-CAR（早期传代和晚期传代）和HepaRG早期传代的聚类，而不是HepaRG晚期传代的聚类，表明传代对HepaRG转录谱的影响比HepaRG-CAR细胞更大。 总之，HepaRG细胞中CAR的过度表达改善了其胆汁酸产生、线粒体能量代谢和稳定性，后者可能是由于ROS产生减少，从而优化了人肝细胞的来源。