Expression and function of the urea cycle in widely-used hepatic cellular models
Expression and function of the urea cycle in widely-used hepatic cellular models
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DOI：
10.1002/jimd.12701
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发表时间：
2024-01-08
影响因子：
4.2
通讯作者：
Haberle，Johannes
中科院分区：
文献类型：
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作者：
Makris，Georgios;Veit，Lara;Haberle，Johannes
一组罕见的代谢缺陷称为尿素循环障碍（UCD）发生在氨消除途径中，并导致失代偿期患者的显著神经认知后遗症。除了原位肝移植，UCD缺乏治疗选择，饮食管理是临床的黄金标准。新的治疗方法对UCD至关重要;然而，这种努力的前提是在有效捕获疾病表现的细胞模型中进行临床前测试。存在几种细胞和动物模型，旨在概括UCD的广泛表型谱;然而，其中大多数缺乏广泛的分子和生化特征。细胞模型的开发正在兴起，因为动物模型非常耗时和成本，并且受到伦理考虑的影响，包括支持动物福利而不是未经检查的临床前试验的3R原则。本研究的目的是比较两种商业肝癌衍生细胞系（诱导多能干细胞肝细胞（iPSC-Hep）、原代人肝细胞（PHH）和人肝细胞制剂）中尿素循环的表达程度和功能。使用尿素循环代谢物的免疫印迹、免疫细胞化学和稳定同位素示踪，我们确定了肝癌衍生的2周分化的HepaRG细胞是尿素循环熟练的，并且表现为PHH的细胞替代物。此外，HepaRG细胞优于iPSC-Heps，已知iPSC-Heps在肝脏成熟度和酶表达方面表现出批次间差异性，因此HepaRG细胞优于iPSC-Heps。上级。最后，HepG 2细胞缺乏尿素循环酶鸟氨酸转氨甲酰酶和尿素酶1，转运蛋白ORNT 1，这限制了它们作为UCD研究模型的适用性。
The group of rare metabolic defects termed urea cycle disorders (UCDs) occur within the ammonia elimination pathway and lead to significant neurocognitive sequelae for patients surviving decompensation episodes. Besides orthotopic liver transplantation, curative options are lacking for UCDs, with dietary management being the gold clinical standard. Novel therapeutic approaches are essential for UCDs; however, such effort presupposes preclinical testing in cellular models that effectively capture disease manifestation. Several cellular and animal models exist and aim to recapitulate the broad phenotypic spectrum of UCDs; however, the majority of those lack extensive molecular and biochemical characterization. The development of cellular models is emerging since animal models are extremely time and cost consuming, and subject to ethical considerations, including the 3R principle that endorses animal welfare over unchecked preclinical testing. The aim of this study was to compare the extent of expression and functionality of the urea cycle in two commercial hepatoma‐derived cell lines, induced pluripotent stem cell hepatocytes (iPSC‐Heps), primary human hepatocytes (PHHs) and human liver cell preparations. Using immunoblotting, immunocytochemistry, and stable isotope tracing of the urea cycle metabolites, we identified that the hepatoma‐derived, 2‐week differentiated HepaRG cells are urea cycle proficient and behave as cellular alternatives to PHHs. Furthermore, HepaRG cells were superior to iPSC‐Heps, which are known to exhibit batch‐to‐batch variabilities in terms of hepatic maturity and enzyme expression. Finally, HepG2 cells lack the urea cycle enzymes ornithine transcarbamylase and arginase 1, the transporter ORNT1, which limits their suitability as model for the study of UCDs.