Comparison of Hepatic 2D Sandwich Cultures and 3D Spheroids for Long-term Toxicity Applications: A Multicenter Study.
Comparison of Hepatic 2D Sandwich Cultures and 3D Spheroids for Long-term Toxicity Applications: A Multicenter Study.
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DOI：
10.1093/toxsci/kfx289
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发表时间：
2018-04-01
期刊：
影响因子：
--
通讯作者：
Ingelman-Sundberg M
中科院分区：
文献类型：
--
作者：
Bell CC;Dankers ACA;Lauschke VM;Sison-Young R;Jenkins R;Rowe C;Goldring CE;Park K;Regan SL;Walker T;Schofield C;Baze A;Foster AJ;Williams DP;van de Ven AWM;Jacobs F;Houdt JV;Lähteenmäki T;Snoeys J;Juhila S;Richert L;Ingelman-Sundberg M
原代人肝细胞（PHH）通常用于药物诱导的肝损伤的体外研究。然而，当作为2D单层培养时，PHH在数小时内失去关键的肝功能。当PHH以三明治构型（2Dsw）培养时，特别是当培养物定期用细胞外基质重新覆盖或作为3D球状体时，可以改善这种去分化。在本研究中，考虑到供体间和实验室间变异性，6个参与实验室评价了由来自相同供体的冻存PHH制成的这2个模型系统的耐用性，并比较了其用于使用5种不同毒性机制的不同肝毒素的重复给药毒性研究的适用性。我们发现，参与药物吸收、分布、代谢和排泄的蛋白质的表达水平，以及5种不同CYP的催化活性，在3D球体培养物中显著更高，可能影响细胞对药物及其代谢物的暴露。此外，全球蛋白质组学分析显示，PHH在3D球状体配置是暂时稳定的，而来自相同的捐助者在2Dsw文化的蛋白质组显示在14天的文化中的蛋白质表达模式的实质性改变。总体而言，球状体培养物对研究的肝毒性化合物更敏感，特别是在长期暴露时，在各试验中心之间几乎没有实验室间或供体间变异性。本文提供的数据表明，重复给药方案提高了体外毒性试验的预测性，并且PHH球状体为药物诱导的肝毒性的长期机制研究提供了一个敏感和稳健的系统，而2Dsw系统具有更多的去分化表型和较低的检测肝毒性的灵敏度。
Primary human hepatocytes (PHHs) are commonly used for in vitro studies of drug-induced liver injury. However, when cultured as 2D monolayers, PHH lose crucial hepatic functions within hours. This dedifferentiation can be ameliorated when PHHs are cultured in sandwich configuration (2Dsw), particularly when cultures are regularly re-overlaid with extracellular matrix, or as 3D spheroids. In this study, the 6 participating laboratories evaluated the robustness of these 2 model systems made from cryopreserved PHH from the same donors considering both inter-donor and inter-laboratory variability and compared their suitability for use in repeated-dose toxicity studies using 5 different hepatotoxins with different toxicity mechanisms. We found that expression levels of proteins involved in drug absorption, distribution, metabolism, and excretion, as well as catalytic activities of 5 different CYPs, were significantly higher in 3D spheroid cultures, potentially affecting the exposure of the cells to drugs and their metabolites. Furthermore, global proteomic analyses revealed that PHH in 3D spheroid configuration were temporally stable whereas proteomes from the same donors in 2Dsw cultures showed substantial alterations in protein expression patterns over the 14 days in culture. Overall, spheroid cultures were more sensitive to the hepatotoxic compounds investigated, particularly upon long-term exposures, across testing sites with little inter-laboratory or inter-donor variability. The data presented here suggest that repeated-dosing regimens improve the predictivity of in vitro toxicity assays, and that PHH spheroids provide a sensitive and robust system for long-term mechanistic studies of drug-induced hepatotoxicity, whereas the 2Dsw system has a more dedifferentiated phenotype and lower sensitivity to detect hepatotoxicity.
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影响因子：
120.1
作者：
Cook, David;Brown, Dearg;Pangalos, Menelas N.
通讯作者：
Pangalos, Menelas N.
影响因子：
3.2
作者：
den Braver-Sewradj, Shalenie P.;den Braver, Michiel W.;Vos, J. Chris
通讯作者：
Vos, J. Chris
影响因子：
6.1
作者：
Heslop JA;Rowe C;Walsh J;Sison-Young R;Jenkins R;Kamalian L;Kia R;Hay D;Jones RP;Malik HZ;Fenwick S;Chadwick AE;Mills J;Kitteringham NR;Goldring CE;Kevin Park B
通讯作者：
Kevin Park B
影响因子：
3.9
作者：
Burbank, Matthew G.;Burban, Audrey;Guillouzo, Andre
通讯作者：
Guillouzo, Andre
影响因子：
13.5
作者：
Lauschke, Volker M.;Vorrink, Sabine U.;Ingelman-Sundberg, Magnus
通讯作者：
Ingelman-Sundberg, Magnus