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J Appl Toxicol. 2015 Aug;35(8):909-17. doi: 10.1002/jat.3090. Epub 2014 Dec 15.

HepaRG culture in tethered spheroids as an in vitro three-dimensional model for drug safety screening.

Wang Z1,2, Luo X1, Anene-Nzelu C3, Yu Y4, Hong X4, Singh NH5, Xia L4, Liu S1, Yu H3,4,5,6,7,8,9,10.

Author information

1
Department of Gastroenterology, Nanfang Hospital, Southern Medical University, No. 1838, North of Guangzhou Dadao, Guangzhou, 510515, China.
2
Department of Gastroenterology, Beijing Chao-Yang Hospital, Capital Medical University, No. 8, South Road of Workers' Gymnasium, Beijing, 100020, China.
3
Department of Bioengineering, National University of Singapore, Block EA, #03-12, 9 Engineering Drive 1, Singapore, 117576, Singapore.
4
Department of Physiology, Yong Loo Lin School of Medicine, National University of Singapore, 10 Medical Drive, Clinical Research Center, #04-25, Singapore, 117597, Singapore.
5
Institute of Bioengineering and Nanotechnology, A*STAR, The Nanos, #04-01, 31 Biopolis Way, Singapore, 138669, Singapore.
6
NUS Graduate School for Integrative Sciences and Engineering, Centre for Life Sciences (CeLS), #05-01, 28 Medical Drive, Singapore, 117576, Singapore.
7
Singapore-MIT Alliance, Computational and System Biology Program, E4-04-10, 4 Engineering Drive 3, Singapore, 117576, Singapore.
8
NUS Tissue Engineering Program, DSO Labs, National University of Singapore, Singapore, 117597, Singapore.
9
Singapore-MIT Alliance for Research and Technology, 3 Science Drive 2, S16-05-08, Singapore, 117543, Singapore.
10
Department of Biological Engineering, Massachusetts Institute of Technology, 77 Massachusetts Ave., Cambridge, MA, 02139, USA.

Abstract

Conventional two-dimensional (2D) monolayer cultures of HepaRG cells allow in vitro maintenance of many liver-specific functions. However, cellular dedifferentiation and functional deterioration over an extended culture period in the conventional 2D HepaRG culture have hampered its applications in drug testing. To address this issue, we developed tethered spheroids of HepaRG cells on Arg-Gly-Asp (RGD) and galactose-conjugated substratum with an optimized hybrid ratio as an in vitro three-dimensional (3D) human hepatocyte model. The liver-specific gene expression level and drug metabolizing enzyme activities in HepaRG-tethered spheorids were markedly higher than those in 2D cultures throughout the culture period of 7 days. The inducibility of three major cytochrome P450 (CYP) enzymes, namely CYP1A2, CYP2B6 and CYP3A4, was improved in both mRNA and activity level in tethered spheroids. Drug-induced cytotoxic responses to model hepatotoxins (acetaminophen, chlorpromazine and ketoconazole) in tethered spheroids were comparable to 2D cultures as well as other studies in the literature. Our results suggested that the HepaRG-tethered spheroid would be an alternative in vitro model suitable for drug safety screening.

KEYWORDS:

3D in vitro model; CYP induction; HepaRG; RGD; drug safety testing; galactose; xenobiotics

PMID:
25512232
DOI:
10.1002/jat.3090
[Indexed for MEDLINE]

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