期刊
TISSUE ENGINEERING PART C-METHODS
卷 24, 期 9, 页码 534-545出版社
MARY ANN LIEBERT, INC
DOI: 10.1089/ten.tec.2018.0134
关键词
3D spheroids; coculture; nonparenchymal cells; primary human hepatocytes
资金
- European Community under Innovative Medicines Initiative (IMI)-Programme (MIP-DILI) [115336]
- European Union [ETB-2015-113]
- European Regional Development Funding (ERDF) [AL0006907]
- Grand Est Region (France)
- Eurostars Programme [E! 11034]
Recent advances in the development of various culture platforms are promising for achieving more physiologically relevant in vitro hepatic models using primary human hepatocytes (PHHs). Previous studies have shown the value of PHHs three-dimensional (3D) spheroid models, cultured in low cell number (1330-2000 cells/3D spheroid), to study long-term liver function as well as pharmacological drug effects and toxicity. In this study, we report that only plateable PHHs aggregate and form compact 3D spheroids with a success rate of 79%, and 96% reproducibility. Out of 3D spheroid forming PHH lots, 65% were considered stable (<50% ATP decrease) over the subsequent 14 days of culture, with reproducibility of a given PHH lot being 82%. We also report successful coculturing of PHHs with human liver nonparenchymal cells (NPCs). Crude P1(c)-NPC fractions were obtained by low centrifugation of the PHH supernatant fraction followed by a few days of culture before harvesting and cryopreservation. At aggregation of PHHs/P1(c)-NPCs (2:1 ratio 3D spheroids), liver sinusoidal endothelial cells, Kupffer cells, and hepatic stellate cells were successfully integrated and remained present throughout the subsequent 14-day culture period as revealed by mRNA expression markers and immunostaining. Increased mRNA expression of albumin (ALB), apolipoprotein B (APOB), cytochrome P450 3A4 (CYP3A4), and increased albumin secretion compared to PHH 3D spheroid monocultures highlighted that in a 3D spheroid coculture, configuration with NPCs, PHH functionality is increased. We thus achieved the development of a more integrated coculture model system requiring low cell numbers, of particular interest due to the scarcity of human liver NPCs.
作者
我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。
推荐
暂无数据