Serum-free collagen sandwich cultures of adult rat hepatocytes maintain liver-like properties long term: A valuable model for in vitro toxicity and drug–drug interaction studies

Tuschl, Gregor; Hrach, Jens; Walter, Yvonne; Hewitt, Philip; Mueller, Stefan O.

doi:10.1016/j.cbi.2009.05.015

Cited by 84 publications

(78 citation statements)

References 48 publications

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“…Other authors confirmed the same trend, leading to prolonged cell differentiation (Tuschl et al, 2009). Vollmar and Menger (2009), in a very recent review also pointed out the role of microcirculation to guarantee hepatic homeostasis: indeed, not only is a sufficient nutritive perfusion and oxygen supply required, but also a balanced vasomotor control and appropriate cell-cell communication.…”

Section: Introductionmentioning

confidence: 76%

Impact of alginate type and bead diameter on mass transfers and the metabolic activities of encapsulated C3A cells in bioartificial liver applications

Gautier

Carpentier²,

Dufresne³

et al. 2011

eCM

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Liver-assist devices have been developed in the last few decades to support patients with liver failure on the road to recovery or transplantation. Fluidised bed bio-artificial livers -where liver cells are encapsulated within alginate beads -appear to be a valuable alternative to hollow fibre devices for improving mass transfers and enhancing treatment efficacy. This approach nevertheless deserves optimization in terms of bead production. The aim of this study was to investigate the impact of alginate type and of two bead diameters (1000 μm and 600 μm) on mass transfers within beads and on the biological functions of encapsulated C3A cells.After assessing the effect of the encapsulation process on bead quality, we investigated cell viability and metabolic activities (ammonia, albumin, α-foetoprotein synthesis and glucose consumption). They were successfully maintained over 48 h within fluidised bed bioreactors, independently of alginate type and bead diameter. Mass transfers were not significantly influenced by the latter parameters. Finally, suggestions are made for improving the entrapment process as a means of enhancing the treatment efficiency of the fluidised bed bioartificial liver.

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Section: Introductionmentioning

confidence: 76%

Impact of alginate type and bead diameter on mass transfers and the metabolic activities of encapsulated C3A cells in bioartificial liver applications

Gautier

Carpentier²,

Dufresne³

et al. 2011

eCM

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“…In our previous study, we showed the serum-free cultivation of PHH in these 3D bioreactors for 11 days. Serum-free cultivation of PHH is of particular interest since it was reported that FCS induces dedifferentiation and therefore loss of functionality of these cells (Tuschl et al 2009). …”

Section: Discussionmentioning

confidence: 99%

Real-time in situ viability assessment in a 3D bioreactor with liver cells using resazurin assay

et al. 2012

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Three-dimensional cultivation of human cells is promising especially for long-term maintenance of specific functions and mimicking the in vivo tissue environment. However, direct viability assessment is very difficult in such systems. Commonly applied indirect methods such as glucose consumption, albumin or urea production are greatly affected by culture conditions, stress and time of cultivation and do not reflect the real time viability of the cells. In this study we established a real-time in situ viability assay namely; resazurin assay, in a 3D hollow-fiber bioreactor using human liver cells. Resazurin assay is based on the conversion of resazurin to a fluorescent dye by cytoplasmatic and mitochondrial enzymes. We show that the resazurin reagent in concentrations used in this study is non-toxic and could be rapidly removed out of the system. Moreover, we observed that dead cells do not affect the results of the assay. We optimized the assay on HepG2 cells and tested it with primary human hepatocytes. Moreover, we maintained primary human hepatocytes in the 3D bioreactor system in serum-free conditions and also assessed viability before and after the exposure to amiodarone using the resazurin assay. We show that this approach is applicable during longterm cultivation of cells in bioreactors under different conditions and can moreover be applied to pharmacological studies, e.g. investigation of chronic drug effects in such 3D bioreactors.

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“…The expression of many liver-specific genes decreases during the initial stage of cultivation of PHHs (Boess et al, 2003;Richert et al, 2006), often associated with declines in metabolic capacities (Richert et al, 2002;Binda et al, 2003;Madan et al, 2003). They are, however, maintained at acceptable levels over time in longterm cultures using sandwich configuration and an appropriate medium (Hewitt et al, 2007;Tuschl et al, 2009;Mueller et al, 2012).…”

Section: Introductionmentioning

confidence: 99%

Transcriptomic Hepatotoxicity Signature of Chlorpromazine after Short- and Long-Term Exposure in Primary Human Sandwich Cultures

Parmentier

Truisi

Moenks

et al. 2013

Drug Metabolism and Disposition

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Drug-induced liver injury is the most frequent reason for market withdrawal of approved drugs, and is difficult to predict in animal models. Here, we analyzed transcriptomic data derived from shortand long-term cultured primary human hepatocytes (PHH) exposed to the well known human hepatotoxin chlorpromazine (CPZ). Samples were collected from five PHH cultures after short-term (1 and 3 days) and long-term (14 days) repeat daily treatment with 0.1 or 0.2 mM CPZ, corresponding to C max . Two PHH cultures were additionally treated with 1 mM CPZ, and the three others with 0.02 mM CPZ. Differences in the total number of gene changes were seen between donors and throughout treatment. Specific transcriptomic hepatotoxicity signatures were created for CPZ and consisted of inflammation/hepatitis, cholestasis, and liver proliferation in all five donors, as well as fibrosis and steatosis, which were observed in four of five donors. Necrosis was present in three of five donors, and an indicative signature of cirrhosis was observed after long-term 14-day repeat treatment, also in three of five donors. The inter-donor variability in the inflammatory response to CPZ treatment was associated with variability in the strength of the response of the transcriptomic hepatotoxicity signatures, suggesting that features of inflammation could be related to the idiosyncratic hepatotoxic effects of CPZ in humans.

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Serum-free collagen sandwich cultures of adult rat hepatocytes maintain liver-like properties long term: A valuable model for in vitro toxicity and drug–drug interaction studies

Cited by 84 publications

References 48 publications

Impact of alginate type and bead diameter on mass transfers and the metabolic activities of encapsulated C3A cells in bioartificial liver applications

Impact of alginate type and bead diameter on mass transfers and the metabolic activities of encapsulated C3A cells in bioartificial liver applications

Real-time in situ viability assessment in a 3D bioreactor with liver cells using resazurin assay

Transcriptomic Hepatotoxicity Signature of Chlorpromazine after Short- and Long-Term Exposure in Primary Human Sandwich Cultures

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