Comparative transcriptomic and phenotypic analysis of induced pluripotent stem cell hepatocyte-like cells and primary human hepatocytes
Comparative transcriptomic and phenotypic analysis of induced pluripotent stem cell hepatocyte-like cells and primary human hepatocytes
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DOI：
10.1007/s00441-024-03868-9
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发表时间：
2024-02-19
影响因子：
3.6
通讯作者：
Rajagopalan，Padmavathy
中科院分区：
文献类型：
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作者：
Gandhi，Neeti;Wills，Lauren;Rajagopalan，Padmavathy
原代人肝细胞（PHH）广泛用于体外肝脏培养以研究肝功能。然而，有限的可用性和侵入性检索阻碍了它们的广泛使用。诱导多能干细胞表现出巨大的潜力，因为它们可以非侵入性地获得并分化成肝谱系，例如肝细胞样细胞（iHLC）。然而，与培养中的 PHH 相比，其胎儿表型特征及其肝功能令人担忧。因此，我们进行了 RNA 测序 (RNA-seq) 分析，以了解每种细胞类型上调或下调的途径。 RNA-seq 数据分析显示胆汁分泌途径上调，其中 AQP9 和 UGT1A1 等基因在 PHH 中的表达量比 iHLC 分别高 455 倍和 15 倍。免疫染色显示，PHH 中存在胆小管。与 iHLC 相比，PHH 中的 TCA 循环上调。细胞分析显示，与 iHLC 相比，PHH 的标准化尿素产量增加了 2-2.5 倍。此外，与 iHLC 相比，PHH 中的药物代谢途径，包括细胞色素 P450 (CYP450) 和 UDP-葡萄糖醛酸基转移酶，均上调。值得注意的是，PHH 中的 CYP2E1 基因表达显着较高（21,810 倍）。对 PHH 和 iHLC 培养物施用对乙酰氨基酚和乙醇，以研究生物转化的差异。与 iHLC 相比，PHH 中基线和毒物处理样品的 CYP450 活性显着更高。我们的分析表明，iHLC 与 PHH 在关键肝功能方面存在显着差异。这些结果强调了 PHH 和 iHLC 之间基因表达和肝功能的差异，以激发未来的研究。图形摘要使用 RNA 测序和表型测量来表征原代人肝细胞 (PHH) 和诱导多能干细胞肝细胞样细胞 (iHLC) 之间的转录组差异
Primary human hepatocytes (PHHs) are used extensively for in vitro liver cultures to study hepatic functions. However, limited availability and invasive retrieval prevent their widespread use. Induced pluripotent stem cells exhibit significant potential since they can be obtained non-invasively and differentiated into hepatic lineages, such as hepatocyte-like cells (iHLCs). However, there are concerns about their fetal phenotypic characteristics and their hepatic functions compared to PHHs in culture. Therefore, we performed an RNA-sequencing (RNA-seq) analysis to understand pathways that are either up- or downregulated in each cell type. Analysis of the RNA-seq data showed an upregulation in the bile secretion pathway where genes such as AQP9 and UGT1A1 were higher expressed in PHHs compared to iHLCs by 455- and 15-fold, respectively. Upon immunostaining, bile canaliculi were shown to be present in PHHs. The TCA cycle in PHHs was upregulated compared to iHLCs. Cellular analysis showed a 2–2.5-fold increase in normalized urea production in PHHs compared to iHLCs. In addition, drug metabolism pathways, including cytochrome P450 (CYP450) and UDP-glucuronosyltransferase enzymes, were upregulated in PHHs compared to iHLCs. Of note, CYP2E1 gene expression was significantly higher (21,810-fold) in PHHs. Acetaminophen and ethanol were administered to PHH and iHLC cultures to investigate differences in biotransformation. CYP450 activity of baseline and toxicant-treated samples was significantly higher in PHHs compared to iHLCs. Our analysis revealed that iHLCs have substantial differences from PHHs in critical hepatic functions. These results have highlighted the differences in gene expression and hepatic functions between PHHs and iHLCs to motivate future investigation.Graphical AbstractCharacterizing the transcriptomic differences between primary human hepatocytes (PHHs) and induced pluripotent stem cell hepatocyte-like cells (iHLCs) using RNA-sequencing and phenotypic measurements