Maintenance of liver functions in rat hepatocytes cultured as spheroids in a rotating wall vessel

Brown, Lanika A.; Arterburn, Linda M.; Miller, Ana P.; Cowger, Nancy L.; Hartley, Sonya M.; Andrews, Annette; Silber, Paul M.; Li, Albert P.

doi:10.1290/020604

Cited by 7 publications

(10 citation statements)

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“…The NASA rotary system, a milestone in dynamic 3D culturing, is a rotating cell culture vessel that simulates a microgravity condition. The low shear force allows spheroid growth as well as high mass transfer of nutrients in media preventing cell death within the spheroid core ( Brown et al, 2003 ). This system has been used to culture spheroids of primary hepatocytes, presenting functional bile canaliculi, up-regulation of hepatocyte-specific functional genes, glycogen storage, as well as ALB production and phase I and II enzymatic activity ( Brown et al, 2003 ; Nelson et al, 2010 ; Chang and Hughes-Fulford, 2014 ).…”

Section: Liver In Vitro Models For Toxicological Smentioning

confidence: 99%

A Critical Perspective on 3D Liver Models for Drug Metabolism and Toxicology Studies

Serras

Rodrigues

Cipriano

et al. 2021

Front. Cell Dev. Biol.

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The poor predictability of human liver toxicity is still causing high attrition rates of drug candidates in the pharmaceutical industry at the non-clinical, clinical, and post-marketing authorization stages. This is in part caused by animal models that fail to predict various human adverse drug reactions (ADRs), resulting in undetected hepatotoxicity at the non-clinical phase of drug development. In an effort to increase the prediction of human hepatotoxicity, different approaches to enhance the physiological relevance of hepatic in vitro systems are being pursued. Three-dimensional (3D) or microfluidic technologies allow to better recapitulate hepatocyte organization and cell-matrix contacts, to include additional cell types, to incorporate fluid flow and to create gradients of oxygen and nutrients, which have led to improved differentiated cell phenotype and functionality. This comprehensive review addresses the drug-induced hepatotoxicity mechanisms and the currently available 3D liver in vitro models, their characteristics, as well as their advantages and limitations for human hepatotoxicity assessment. In addition, since toxic responses are greatly dependent on the culture model, a comparative analysis of the toxicity studies performed using two-dimensional (2D) and 3D in vitro strategies with recognized hepatotoxic compounds, such as paracetamol, diclofenac, and troglitazone is performed, further highlighting the need for harmonization of the respective characterization methods. Finally, taking a step forward, we propose a roadmap for the assessment of drugs hepatotoxicity based on fully characterized fit-for-purpose in vitro models, taking advantage of the best of each model, which will ultimately contribute to more informed decision-making in the drug development and risk assessment fields.

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Section: Liver In Vitro Models For Toxicological Smentioning

confidence: 99%

A Critical Perspective on 3D Liver Models for Drug Metabolism and Toxicology Studies

Serras

Rodrigues

Cipriano

et al. 2021

Front. Cell Dev. Biol.

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“…Higher recovery rates of viable cells were found in suspension groups (46.6%) compared with the trypsinized adhesion group (37.1%). However, based on encouraging results from long‐term extracellular matrix (ECM)‐free cultures in the RCCS of murine, porcine, and human hepatocytes (22–24), we expected higher recovery rates. Moreover, viability was lower in the suspension‐labeled group compared with the trypsinized cells (47.8 ± 2.8% vs. 60.9 ± 4.5%).…”

Section: Discussionmentioning

confidence: 99%

“…Bioreactors are available with increasing volumes, from 10 to 500 mL. Several studies report promising results regarding long‐term survival and prolonged cell performance of hepatic tumor cell lines and murine, porcine, and human hepatocytes in suspension culture (21–24). Taking into account recent developments in the field of hepatocyte transplantation, the aim of this study was to establish a protocol for fast, easy, and clinically applicable labeling of PHHs with MPIOs in suspension for routine large‐scale labeling procedures, with the potential for future GMP‐conforming processing.…”

mentioning

confidence: 99%

Labeling of Primary Human Hepatocytes With Micron‐Sized Iron Oxide Particles in Suspension Culture Suitable for Large‐Scale Preparation

et al. 2011

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Labeling of hepatocytes with micron-sized iron oxide particles (MPIOs) enables cell detection using clinical magnetic resonance equipment. For clinical applications, large numbers of cells must be labeled in a simple and rapid manner and have to be applied in suspension. However, all existing protocols are based on adhesion culture labeling with subsequent resuspension, only suitable for small experimental settings. The aim of this study was to investigate the feasibility of preparing MPIO-labeled primary human hepatocytes in a temporary suspension culture. Human hepatocytes were isolated from 16 donors and labeled with MPIOs in suspension, using the Rotary Cell Culture System. Particle incorporation was investigated by light and electron microscopy. Cells were compared with adhesion culture-labeled and subsequently enzymatically resuspended cells. During a period of 5 days, hepatocyte-specific parameters of cell damage (aspartate aminotransferase and alanine aminotransferase) and metabolic activity (urea and albumin) were analyzed (n=7). Suspension cultures showed a higher outcome in cell recovery compared with the conventional labeling method. When incubated with 180 particles/viable cell for 4 h, the mean particle uptake was 28.8 particles/cell at a labeling efficiency of 95.1%. Labeling in suspension had no adverse effects on cell integrity or metabolic activity. We conclude that labeling of human hepatocytes in suspension is feasible and simple and may serve future large-scale processing of cells.

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“…Hepatocyte spheroids have been produced by (Brown et al, 2003) and maintained in a differentiated state for several weeks by culturing in a rotating well vessel (RWV, www.synthecon.com). The spheroids were obtained by culturing primary rat hepatocytes in 100-mm Petri dishes constantly shacked with an orbital shaker.…”

Section: Spheroid-based Liver Modelsmentioning

confidence: 99%

Three-Dimensional Cell Cultures in Toxicology

Pampaloni

Stelzer

2009

Biotechnology and Genetic Engineering Reviews

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Toxicity testing with animals is expensive, ethically controversial, and not always predictive of the human response. Cell-based assays are regarded as an alternative. However, conventional two-dimensional cell cultures do not reproduce the tissue architecture in vivo, and do not forecast organ-specific toxicity. On the other hand, three-dimensional cultures emulate the biochemistry and mechanics of the microenvironment in tissues more closely. Therefore, they address the limitations of both animals and two-dimensional cultures, and provide more accurate data on the effects of short-and long-term exposure to toxicants. We provide an up-to-date overview on the use of three-dimensional cell cultures in toxicology. We anticipate that threedimensional cultures will become invaluable to accomplish the 3R agenda (refinement, reduction, and replacement) for animal-based toxicity testing and will play a major role for the Registration, Evaluation and Authorisation of Chemicals in the European Union (REACH legislation).

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Maintenance of liver functions in rat hepatocytes cultured as spheroids in a rotating wall vessel

Cited by 7 publications

References 0 publications

A Critical Perspective on 3D Liver Models for Drug Metabolism and Toxicology Studies

A Critical Perspective on 3D Liver Models for Drug Metabolism and Toxicology Studies

Labeling of Primary Human Hepatocytes With Micron‐Sized Iron Oxide Particles in Suspension Culture Suitable for Large‐Scale Preparation

Three-Dimensional Cell Cultures in Toxicology

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