Rat Primary Hepatocytes Show Enhanced Performance and Sensitivity to Acetaminophen During Three-Dimensional Culture on a Polystyrene Scaffold Designed for Routine Use

Schutte, Maaike E.; Fox, Bridget; Baradez, Marc‐Olivier; Devonshire, Alison S.; Minguez, Jesus; Bokhari, Maria; Przyborski, Stefan; Marshall, Damian

doi:10.1089/adt.2011.0371

Cited by 68 publications

(45 citation statements)

References 45 publications

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“…5c), we quantified the proportion of viable microtissues when treated with increasing APAP doses and observed a nonlinear toxicity ''shoulder'' below which there was little effect, but above which hepatotoxicity is observed. The shape of this dose curve may be attributed to the depletion of intracellular glutathione 52 and has been observed in other primary hepatocyte systems, 39,53 indicating that these microtissues faithfully detect a canonical example of primary hepatocyte toxicity.…”

Section: Characterization Of Microtissue Drug Metabolism Enzyme Activitymentioning

confidence: 52%

Micropatterned Cell–Cell Interactions Enable Functional Encapsulation of Primary Hepatocytes in Hydrogel Microtissues

Stevens

Schwartz

et al. 2014

Tissue Engineering Part A

123

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Drug-induced liver injury is a major cause of drug development failures and postmarket withdrawals. In vitro models that incorporate primary hepatocytes have been shown to be more predictive than model systems which rely on liver microsomes or hepatocellular carcinoma cell lines. Methods to phenotypically stabilize primary hepatocytes ex vivo often rely on mimicry of hepatic microenvironmental cues such as cell-cell interactions and cell-matrix interactions. In this work, we sought to incorporate phenotypically stable hepatocytes into threedimensional (3D) microtissues, which, in turn, could be deployed in drug-screening platforms such as multiwell plates and diverse organ-on-a-chip devices. We first utilize micropatterning on collagen I to specify cell-cell interactions in two-dimensions, followed by collagenase digestion to produce well-controlled aggregates for 3D encapsulation in polyethylene glycol (PEG) diacrylate. Using this approach, we examined the influence of homotypic hepatocyte interactions and composition of the encapsulating hydrogel, and achieved the maintenance of liver-specific function for over 50 days. Optimally preaggregated structures were subsequently encapsulated using a microfluidic droplet-generator to produce 3D microtissues. Interactions of engineered hepatic microtissues with drugs was characterized by flow cytometry, and yielded both induction of P450 enzymes in response to prototypic small molecules and drug-drug interactions that give rise to hepatotoxicity. Collectively, this study establishes a pipeline for the manufacturing of 3D hepatic microtissues that exhibit stabilized liver-specific functions and can be incorporated into a wide array of emerging drug development platforms.

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Section: Characterization Of Microtissue Drug Metabolism Enzyme Activitymentioning

confidence: 52%

Micropatterned Cell–Cell Interactions Enable Functional Encapsulation of Primary Hepatocytes in Hydrogel Microtissues

Stevens

Schwartz

et al. 2014

Tissue Engineering Part A

123

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“…The results for viability in Figure 6 are in agreement with a number of published studies, where survival is enhanced in sandwhich culture. [34][35][36][37] …”

Section: Survival Of Primary Rat Hepatocyte (Prh) On Substrates With mentioning

confidence: 99%

Primary Liver Cells Cultured on Carbon Nanotube Substrates for Liver Tissue Engineering and Drug Discovery Applications

Abdullah

Azad

Ovalle‐Robles

et al. 2014

ACS Appl. Mater. Interfaces

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Here we explore the use of two-and three-dimensional scaffolds of multi-walled-carbon nanotubes (MWNTs) for hepatocyte cell culture. Our objective is to study the use of these scaffolds in liver tissue engineering and drug discovery. In our experiments, primary rat hepatocytes, the parenchymal (main functional) cell type in the liver, were cultured on aligned nanogrooved MWNT sheets, MWNT yarns, or standard 2-dimensional culture conditions as a control. We find comparable cell viability between all three culture conditions, but enhanced production of the hepatocyte-specific marker albumin for cells cultured on MWNTs. The basal activity of two clinically relevant cytochrome P450 enzymes, CYP1A2 and CYP3A4, are similar on all substrates, but we find enhanced induction of CYP1A2 for cells on the MWNT sheets. Our data thus supports the use of these substrates for applications including tissue engineering, enhancing liver-specific functions, as well an in vitro model systems with enhanced predictive capability in drug discovery and development.

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“…For example rat primary hepatocytes show a more cuboidal morphology, increased viability, greater induction of cytochrome-P450 enzyme activity and increased sensitivity to paracetamol toxicity when grown in a commercially available polystyrene scaffold compared to cells grown in 2D culture 17 . Using the same scaffold the hepatocarcinoma cell line HepG2 has also been shown to increase albumin production 18 and show a more in vivo-like response to methotrexate compared to 2D cultured cells 19 .…”

Section: Introductionmentioning

confidence: 99%

Hollow Fiber Bioreactors for <em>In Vivo</em>-like Mammalian Tissue Culture

Storm

Sorrell

Shipley

et al. 2016

JoVE

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Tissue culture has been used for over 100 years to study cells and responses ex vivo. The convention of this technique is the growth of anchorage dependent cells on the 2-dimensional surface of tissue culture plastic. More recently, there is a growing body of data demonstrating more in vivo-like behaviors of cells grown in 3-dimensional culture systems. This manuscript describes in detail the set-up and operation of a hollow fiber bioreactor system for the in vivo-like culture of mammalian cells. The hollow fiber bioreactor system delivers media to the cells in a manner akin to the delivery of blood through the capillary networks in vivo. The system is designed to fit onto the shelf of a standard CO 2 incubator and is simple enough to be set-up by any competent cell biologist with a good understanding of aseptic technique. The systems utility is demonstrated by culturing the hepatocarcinoma cell line HepG2/C3A for 7 days. Further to this and in line with other published reports on the functionality of cells grown in 3-dimensional culture systems the cells are shown to possess increased albumin production (an important hepatic function) when compared to standard 2-dimensional tissue culture. Video LinkThe video component of this article can be found at

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Rat Primary Hepatocytes Show Enhanced Performance and Sensitivity to Acetaminophen During Three-Dimensional Culture on a Polystyrene Scaffold Designed for Routine Use

Cited by 68 publications

References 45 publications

Micropatterned Cell–Cell Interactions Enable Functional Encapsulation of Primary Hepatocytes in Hydrogel Microtissues

Micropatterned Cell–Cell Interactions Enable Functional Encapsulation of Primary Hepatocytes in Hydrogel Microtissues

Primary Liver Cells Cultured on Carbon Nanotube Substrates for Liver Tissue Engineering and Drug Discovery Applications

Hollow Fiber Bioreactors for <em>In Vivo</em>-like Mammalian Tissue Culture

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