期刊
TOXICOLOGY IN VITRO
卷 55, 期 -, 页码 160-172出版社
PERGAMON-ELSEVIER SCIENCE LTD
DOI: 10.1016/j.tiv.2018.12.014
关键词
Liver spheroids; 3D cell culture; Primary rat hepatocytes; Polarisation; Bile canaliculi; Liquid-overlay technique
类别
资金
- Syngenta Ltd, UK
- Liverpool Centre for Mathematics in Healthcare (EPSRC) [EP/N014499/1]
- Biotechnology and Biological Science Research Council (BBSRC) industrial CASE partnership award [BB/M503435/1]
- EPSRC [EP/N014499/1] Funding Source: UKRI
- MRC [MR/L006758/1] Funding Source: UKRI
Many in vitro liver cell models, such as 2D systems, that are used to assess the hepatotoxic potential of xenobiotics suffer major limitations arising from a lack of preservation of physiological phenotype and metabolic competence. To circumvent some of these limitations there has been increased focus on producing more representative 3D models. Here we have used a novel approach to construct a size-controllable 3D hepatic spheroid model using freshly isolated primary rat hepatocytes (PRH) utilising the liquid-overlay technique whereby PRH spontaneously self-assemble in to 3D microtissues. This system produces viable spheroids with a compact in vivo-like structure for up to 21 days with sustained albumin production for the duration of the culture period. F-actin was seen throughout the spheroid body and P-glycoprotein (P-gp) and multidrug resistance-associated protein 2 (MRP2) transporters had polarised expression on the canalicular membrane of hepatocytes within the spheroids upon formation (day 3). The MRP2 transporter was able to functionally transport 5-mu M 5-chloromethylfluorescein diacetate (CMFDA) substrates into these canalicular structures. These PRH spheroids display in vivo characteristics including direct cell-cell contacts, cellular polarisation, 3D cellular morphology, and formation of functional secondary structures throughout the spheroid. Such a well-characterised system could be readily exploited for pre-clinical and non-clinical repeat-dose investigations and could make a significant contribution to replace, reduce and refine the use of animals for applied research.
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